Controlled release aural pressure modulator compositions and methods for the treatment of OTIC disorders

ABSTRACT

Disclosed herein are compositions and methods for the treatment of otic diseases or conditions with aural pressure modulating agent compositions and formulations administered locally to an individual afflicted with an otic disease or condition, through direct application of these compositions and formulations onto or via perfusion into the targeted auris structure(s).

CROSS-REFERENCE

This application claims the benefit of U.S. Provisional Application No.61/073,716 filed Jun. 18, 2008, U.S. Provisional Application No.61/074,583 filed Jun. 20, 2008, U.S. Provisional Application No.61/082,450 filed Jul. 21, 2008, U.S. Provisional Application No.61/086,105 filed Aug. 4, 2008, U.S. Provisional Application No.61/094,384 filed Sep. 4, 2008, U.S. Provisional Application No.61/101,112 filed Sep. 29, 2008, and U.S. Provisional Application No.61/140,033 filed Dec. 22, 2008, all of which are incorporated herein intheir entirety.

JOINT RESEARCH AGREEMENT

The claimed invention was made as a result of activities undertakenwithin the scope of a joint research agreement between Jay BenjaminLichter, Benedikt K. Vollrath, Otonomy, Inc., and Avalon Ventures VIIIGP, LLC that was in effect on or before the date the invention was made.

BACKGROUND OF THE INVENTION

Vertebrates have a pair of ears, placed symmetrically on opposite sidesof the head. The ear serves as both the sense organ that detects soundand the organ that maintains balance and body position. The ear isgenerally divided into three portions: the outer ear, auris media (ormiddle ear) and the auris interna (or inner ear).

SUMMARY OF THE INVENTION

Described herein are compositions, formulations, manufacturing methods,therapeutic methods, uses, kits, and delivery devices for the controlledrelease or delivery of at least one aural pressure modulating agent toat least one structure or region of the ear.

In some embodiments, the aural pressure modulating agents targetimbalance in aural pressure in the ear. In some embodiments, the auralpressure modulating agents target fluid homeostasis control in the innerear, or auris interna. In other embodiments, the aural pressuremodulating agents target fluid homeostasis control in the middle ear, orauris media. In some embodiments, the aural pressure modulators arevasopressin receptor modulators, prostaglandins or estrogen-relatedreceptor modulators. In some embodiments, the aural pressure modulatingagents are inhibitors of vasopressin receptor function, prostaglandinreceptor or estrogen-related receptor function in the ear. In someembodiments, the aural pressure modulating agents are osmotic diureticagents. In some embodiments, the controlled release formulations furthercomprise a rapid or immediate release component for delivering auralpressure modulating agents to the auris interna. All formulationscomprise excipients that are auris-interna acceptable.

Also disclosed herein are methods and compositions for the treatment ofotic diseases or conditions associated with an imbalance in auralpressure by administration of controlled release formulations comprisingaural pressure modulating agents. In some embodiments, the otic diseaseor condition associated with an imbalance in aural pressure is a fluidhomeostasis disorder. Also disclosed herein is the local delivery ofcontrolled release aural pressure modulating compositions andformulations to suppress or ameliorate auditory and vestibularimpairment as a result of an imbalance in aural pressure and/orimbalance in fluid homeostasis in the ear, such as Meniere's disease. Inanother aspect, the composition is administered so that the compositionis in contact with the crista fenestrae cochleae, the round window orthe tympanic cavity.

Disclosed herein are controlled release formulations for delivering anaural pressure modulating agent to the ear. In some embodiments, thetarget portion of the ear is the middle ear or auris media. In someembodiments, the target portion of the ear is the inner ear, or aurisinterna. In other embodiments, the target portion of the ear is both theauris media and the auris interna.

The auris formulations and therapeutic methods described herein havenumerous advantages that overcome the previously-unrecognizedlimitations of formulations and therapeutic methods described in priorart.

Sterility

The environment of the inner ear is an isolated environment. Theendolymph and the perilymph are static fluids and are not in contiguouscontact with the circulatory system. The blood-labyrinth-barrier (BLB),which includes a blood-endolymph barrier and a blood-perilymph barrier,consists of tight junctions between specialized epithelial cells in thelabyrinth spaces (i.e., the vestibular and cochlear spaces). Thepresence of the BLB limits delivery of active agents (e.g., auralpressure modulators) to the isolated microenvironment of the inner ear.Auris hair cells are bathed in endolymphatic or perilymphatic fluids andcochlear recycling of potassium ions is important for hair cellfunction. When the inner ear is infected, there is an influx ofleukocytes and/or immunoglobins (e.g. in response to a microbialinfection) into the endolymph and/or the perilymph and the delicateionic composition of inner ear fluids is upset by the influx ofleukocytes and/or immunoglobins. In certain instances, a change in theionic composition of inner ear fluids results in hearing loss, loss ofbalance and/or ossification of auditory structures. In certaininstances, even trace amounts of pyrogens and/or microbes can triggerinfections and related physiological changes in the isolatedmicroenvironment of the inner ear.

Due to the susceptibilty of the inner ear to infections, aurisformulations require a level of sterility (e.g., low bioburden) that hasnot been recognized hitherto in prior art. Provided herein are aurisformulations that are manufactured with low bioburden or sterilized withstringent sterility requirements and are suitable for administration tothe middle and/or inner ear. In some embodiments, the auris compatiblecompositions described herein are substantially free of pyrogens and/ormicrobes.

Compatibility with Inner Ear Environment

Described herein are otic formulations with an ionic balance that iscompatible with the perilymph and/or the endolymph and does not causeany change in cochlear potential. In specific embodiments,osmolarity/osmolality of the present formulations is adjusted, forexample, by the use of appropriate salt concentrations (e.g.,concentration of sodium salts) or the use of tonicity agents whichrenders the formulations endolymph-compatible and/orperilymph-compatible (i.e. isotonic with the endolymph and/orperilymph). In some instances, the endolymph-compatible and/orperilymph-compatible formulations described herein cause minimaldisturbance to the environment of the inner ear and cause minimumdiscomfort (e.g, vertigo) to a mammal (e.g., a human) uponadministration. Further, the formulations comprise polymers that arebiodegradable and/or dispersable, and/or otherwise non-toxic to theinner ear environment. In some embodiments, the formulations describedherein are free of preservatives and cause minimal disturbance (e.g.,change in pH or osmolarity, irritation) in auditory structures. In someembodiments, the formulations described herein comprise antioxidantsthat are non-irritating and/or non-toxic to otic structures.

Dosing Frequency

The current standard of care for auris formulations requires multipleadministrations of drops or injections (e.g. intratympanic injections)over several days (e.g., up to two weeks), including schedules ofreceiving multiple injections per day. In some embodiments, aurisformulations described herein are controlled release formulations, andare administered at reduced dosing frequency compared to the currentstandard of care. In certain instances, when an auris formulation isadministered via intratympanic injection, a reduced frequency ofadministration alleviates discomfort caused by multiple intratympanicinjections in individuals undergoing treatment for a middle and/or innerear disease, disorder or condition. In certain instances, a reducedfrequency of administration of intratympanic injections reduces the riskof permanent damage (e.g., perforation) to the ear drum. Theformulations described herein provide a constant, sustained, extended,delayed or pulsatile rate of release of an active agent into the innerear environment and thus avoid any variability in drug exposure intreatment of otic disorders.

Therapeutic Index

Auris formulations described herein are administered into the ear canal,or in the vestibule of the ear. Access to, for example, the vestibularand cochlear apparatus will occur through the auris media including theround window membrane, the oval window/stapes footplate, the annularligament and through the otic capsule/temporal bone. Otic administrationof the formulations described herein avoids toxicity associated withsystemic administration (e.g., hepatotoxicity, cardiotoxicity,gastrointestinal side effects, renal toxicity) of the active agents. Insome instances, localized administration in the ear allows an activeagent to reach a target organ (e.g., inner ear) in the absence ofsystemic accumulation of the active agent. In some instances, localadministration to the ear provides a higher therapeutic index for anactive agent that would otherwise have dose-limiting systemic toxicity.

Prevention of Drainage into Eustachian Tube

In some instances, a disadvantage of liquid formulations is theirpropensity to drip into the eustachian tube and cause rapid clearance ofthe formulation from the inner ear. Provided herein, in certainembodiments, are auris formulations comprising polymers that gel at bodytemperature and remain in contact with the target auditory surfaces(e.g., the round window) for extended periods of time. In someembodiments, the formulations further comprise mucoadhesives that allowthe formulations to adhere to otic mucosal surfaces. In some instances,the auris formulations described herein avoid attenuation of therapeuticbenefit due to drainage or leakage of active agents via the eustachiantube.

Description of Certain Embodiments

Described herein are controlled release compositions and devices fortreating otic disorders comprising a therapeutically-effective amount ofan aural pressure modulating agent, a controlled releaseauris-acceptable excipient and an auris-acceptable vehicle. In oneaspect, the controlled release auris-acceptable excipient is chosen froman auris-acceptable polymer, an auris-acceptable viscosity enhancingagent, an auris-acceptable gel, an auris-acceptable paint, anauris-acceptable foam, an auris-acceptable microsphere or microparticle,an auris-acceptable hydrogel, an auris-acceptable in situ forming spongymaterial, an auris-acceptable actinic radiation curable gel, anauris-acceptable liposome, an auris-acceptable nanocapsule ornanosphere, an auris-acceptable thermoreversible gel or combinationsthereof. In further embodiments, the auris-acceptable viscosityenhancing agent is a cellulose, a cellulose ether, alginate,polyvinylpyrrolidone, a gum, a cellulosic polymer or combinationsthereof. In yet another embodiment, the auris-acceptable viscosityenhancing agent is present in an amount sufficient to provide aviscosity of between about 1000 to about 1,000,000 centipoise. In stillanother aspect, the auris-acceptable viscosity enhancing agent ispresent in an amount sufficient to provide a viscosity of between about50,000 to about 1,000,000 centipoise. In some embodiments, the auralpressure modulating formulations or compositions are optimal forosmolality or osmolarity of the target auris structure to ensurehomeostasis is maintained.

In some embodiments, the compositions are formulated for pH, and apractical osmolality or osmolarity to ensure that homeostasis of thetarget auris structure is maintained. A perilymph-suitableosmolarity/osmolality is a practical/deliverable osmolarity/osmolalitythat maintains the homeostasis of the target auris structure duringadministration of the pharmaceutical formulations described herein.

For example, the osmolarity of the perilymph is between about 270-300mOsm/L, and the compositions described herein are optionally formulatedto provide a practical osmolarity of about 150 to about 1000 mOsm/L. Incertain embodiments, the formulations described herein provide apractical and/or deliverable osmolarity within about 150 to about 500mOsm/L at the target site of action (e.g., the inner ear and/or theperilymph and/or the endolymph). In certain embodiments, theformulations described herein provide a practical osmolarity withinabout 200 to about 400 mOsm/L at the target site of action (e.g., theinner ear and/or the perilymph and/or the endolymph). In certainembodiments, the formulations described herein provide a practicalosmolarity within about 250 to about 320 mOsm/L at the target site ofaction (e.g., the inner ear and/or the perilymph and/or the endolymph).In certain embodiments, the formulations described herein provide aperilymph-suitable osmolarity within about 150 to about 500 mOsm/L,about 200 to about 400 mOsm/L or about 250 to about 320 mOsm/L at thetarget site of action (e.g., the inner ear and/or the perilymph and/orthe endolymph). In certain embodiments, the formulations describedherein provide a perilymph-suitable osmolality within about 150 to about500 mOsm/kg, about 200 to about 400 mOsm/kg or about 250 to about 320mOsm/kg at the target site of action (e.g., the inner ear and/or theperilymph and/or the endolymph). Similarly, the pH of the perilymph isabout 7.2-7.4, and the pH of the present formulations is formulated(e.g., with the use of buffers) to provide a perilymph-suitable pH ofabout 5.5 to about 9.0, about 6.0 to about 8.0 or about 7.0 to about7.6. In certain embodiments, the pH of the formulations is within about6.0 to about 7.6. In certain instances, the pH of the endolymph is about7.2-7.9, and the pH of the present formulations is formulated (e.g.,with the use of buffers) to be within about 5.5 to about 9.0, withinabout 6.5 to about 8.0 or within about 7.0 to about 7.6.

In some aspects, the controlled-release auris-acceptable excipient isbiodegradable. In some aspects the controlled release auris-acceptableexcipient is bioeliminated (e.g., degraded and/or eliminated throughurine, feces or other routes of elimination). In another aspect, thecontrolled release composition further comprises an auris-acceptablemucoadhesive, an auris-acceptable penetration enhancer or anauris-acceptable bioadhesive.

In one aspect, the controlled release aural pressure modulating agentcomposition is delivered using a drug delivery device, which is a needleand syringe, a pump, a microinjection device or combinations thereof. Insome embodiments, the aural pressure modulator of the controlled releasecomposition has limited or no systemic release, is toxic whenadministered systemically, has poor pK characteristics or combinationsthereof. In some aspects, the aural pressure modulator is a smallmolecule agent. In other aspects, the aural pressure modulator is anantibody. In some embodiments, the aural pressure modulator is avasopressin receptor modulator, a prostaglandin receptor modulator or anestrogen-related receptor modulator. In additional aspects, thevasopressin receptor modulator specifically acts on vasopressin receptor2 (VP2).

Also disclosed herein is a method for treating an otic disordercomprising administering at least once every 3, 4, 5, 6, 7, 8, 9, 10,11, 12, 13, 14, or 15 days, at least once a week, once every two weeks,once every three weeks, once every four weeks, once every five weeks, oronce every six weeks; or once a month, once every two months, once everythree months, once every four months, once every five months, once everysix months, once every seven months, once every eight months, once everynine months, once every ten months, once every eleven months, or onceevery twelve months with the compositions and formulations disclosedherein. In particular embodiments, the controlled release formulationsdescribed herein provide a sustained dose of aural pressure modulatingagent to the inner ear between subsequent doses of the controlledrelease formulation. That is, taking one example only, if new doses ofthe aural pressure modulating agent controlled release formulation areadminstered via intratympanic injection to the round window membraneevery 10 days, then the controlled release formulation provides aneffective dose of aural pressure modulating agent to the inner ear(e.g., across the round window membrane) during that 10-day period.

In one aspect, the composition is administered so that the compositionis in contact with the crista fenestrae cochleae, the round windowmembrane or the tympanic cavity. In one aspect the composition isadministered by intratympanic injection.

Provided herein is a pharmaceutical composition or device comprising anamount of an aural pressure modulating agent that is therapeuticallyeffective for treating an otic disease or condition associated with animbalance in aural pressure, the pharmaceutical composition or devicecomprising substantially low degradation products of the auris pressuremodulating agent, the pharmaceutical composition or device furthercomprising two or more characteristics selected from:

-   -   (i) between about 0.1% to about 10% by weight of the aural        pressure modulating agent, or pharmaceutically acceptable        prodrug or salt thereof;    -   (ii) between about 14% to about 21% by weight of a        polyoxyethylene-polyoxypropylene triblock copolymer of general        formula E106 P70 E106;    -   (iii) sterile water, q.s., buffered to provide a pH between        about 5.5 and about 8.0;    -   (iv) multiparticulate aural pressure modulating agent;    -   (v) a gelation temperature between about 19° C. to about 42° C.;    -   (vi) less than about 50 colony forming units (cfu) of        microbiological agents per gram of formulation, and    -   (vii) less than about 5 endotoxin units (EU) per kg of body        weight of a subject.

In some embodiments, a pharmaceutical composition or device describedherein comprises:

-   -   (i) between about 0.1% to about 10% by weight of the aural        pressure modulating agent, or pharmaceutically acceptable        prodrug or salt thereof;    -   (ii) between about 14% to about 21% by weight of a        polyoxyethylene-polyoxypropylene triblock copolymer of general        formula E106 P70 E106; and    -   (iii) multiparticulate aural pressure modulating agent.

In some embodiments, a pharmaceutical composition or device describedherein comprises:

-   -   (i) between about 0.1% to about 10% by weight of the aural        pressure modulating agent, or pharmaceutically acceptable        prodrug or salt thereof;    -   (ii) between about 14% to about 21% by weight of a        polyoxyethylene-polyoxypropylene triblock copolymer of general        formula E106 P70 E106;    -   (iii) multiparticulate aural pressure modulating agent; and    -   (iv) a gelation temperature between about 19° C. to about 42° C.

In some embodiments a pharmaceutical composition or device describedabove provides a practical osmolarity between about 150 and 500 mOsm/L.In some embodiments a pharmaceutical composition or device describedabove provides a practical osmolarity between about 200 and 400 mOsm/L.In some embodiments a pharmaceutical composition or device describedabove provides a practical osmolarity between about 250 and 320 mOsm/L.

In some embodiments, the aural pressure modulating agent is releasedfrom the pharmaceutical composition or device described above for aperiod of at least 3 days. In some embodiments, the aural pressuremodulating agent is released from the pharmaceutical composition ordevice described above for a period of at least 5 days. In someembodiments, the aural pressure modulating agent is released from thepharmaceutical composition or device described above for a period of atleast 10 days. In some embodiments, the aural pressure modulating agentis released from the pharmaceutical composition or device describedabove for a period of at least 14 days. In some embodiments, the auralpressure modulating agent is released from the pharmaceuticalcomposition or device described above for a period of at least onemonth.

In some embodiments, a pharmaceutical composition or device describedabove comprises aural pressure modulating agent as a neutral compound, afree acid, a free base, a salt or a prodrug. In some embodiments, apharmaceutical composition or device described above comprises auralpressure modulating agent as a neutral compound, a free acid, a freebase, a salt or a prodrug, or a combination thereof.

In some embodiments, a pharmaceutical composition or device describedabove is an auris-acceptable thermoreversible gel. In some embodimentsof the pharmaceutical composition or device, thepolyoxyethylene-polyoxypropylene triblock copolymer is bioeliminated.

In some embodiments the pharmaceutical composition or device furthercomprises a penetration enhancer. In some embodiments, thepharmaceutical composition or device further comprises a dye.

In some embodiments of the pharmaceutical composition or devicedescribed above, the aural pressure modulating agent inhibitsvasopressin receptor function, prostaglandin receptor function orestrogen-related receptor beta function, or combinations thereof.

In some embodiments, the pharmaceutical composition or device furthercomprises the aural pressure modulating agent, or pharmaceuticallyacceptable salt thereof, prodrug or combination thereof as an immediaterelease agent.

In some embodiments of the pharmaceutical composition or device theaural pressure modulating agent comprises multiparticulates. In someembodiments of the pharmaceutical composition or device the auralpressure modulating agent is essentially in the form of micronizedparticles. In some embodiments of the pharmaceutical composition ordevice, the aural pressure modulating agent is in the form ofmicro-aural pressure modulating agent powder.

In some embodiments, a pharmaceutical composition or device describedabove comprises aural pressure modulating agent as multiparticulates. Insome embodiments, a pharmaceutical composition or device described abovecomprises aural pressure modulating agent in the form of micronizedparticles. In some embodiments, a pharmaceutical composition or devicedescribed above comprises aural pressure modulating agent as micronizedpowder.

In some embodiments, a pharmaceutical composition or device describedabove comprises about 10% of a polyoxyethylene-polyoxypropylene triblockcopolymer of general formula E106 P70 E106 by weight of the composition.In some embodiments, a pharmaceutical composition or device describedabove comprises about 15% of a polyoxyethylene-polyoxypropylene triblockcopolymer of general formula E106 P70 E106 by weight of the composition.In some embodiments, a pharmaceutical composition or device describedabove comprises about 20% of a polyoxyethylene-polyoxypropylene triblockcopolymer of general formula E106 P70 E106 by weight of the composition.In some embodiments, a pharmaceutical composition or device describedabove comprises about 25% of a polyoxyethylene-polyoxypropylene triblockcopolymer of general formula E106 P70 E106 by weight of the composition.

In some embodiments, a pharmaceutical composition or device describedabove comprises about 0.01% of an aural pressure modulating agent, orpharmaceutically acceptable prodrug or salt thereof, by weight of thecomposition. In some embodiments, a pharmaceutical composition or devicedescribed above comprises about 0.05% of an aural pressure modulatingagent, or pharmaceutically acceptable prodrug or salt thereof, by weightof the composition. In some embodiments, a pharmaceutical composition ordevice described above comprises about 0.1% of an aural pressuremodulating agent, or pharmaceutically acceptable prodrug or saltthereof, by weight of the composition. In some embodiments, apharmaceutical composition or device described above comprises about 1%of an aural pressure modulating agent, or pharmaceutically acceptableprodrug or salt thereof, by weight of the composition. In someembodiments, a pharmaceutical composition or device described abovecomprises about 2.5% of an aural pressure modulating agent, orpharmaceutically acceptable prodrug or salt thereof, by weight of thecomposition. In some embodiments, a pharmaceutical composition or devicedescribed above comprises about 5% of an aural pressure modulatingagent, or pharmaceutically acceptable prodrug or salt thereof, by weightof the composition. In some embodiments, a pharmaceutical composition ordevice described above comprises about 10% of an aural pressuremodulating agent, or pharmaceutically acceptable prodrug or saltthereof, by weight of the composition. In some embodiments, apharmaceutical composition or device described above comprises about 20%of an aural pressure modulating agent, or pharmaceutically acceptableprodrug or salt thereof, by weight of the composition. In someembodiments, a pharmaceutical composition or device described abovecomprises about 30% of an aural pressure modulating agent, orpharmaceutically acceptable prodrug or salt thereof, by weight of thecomposition. In some embodiments, a pharmaceutical composition or devicedescribed above comprises about 40% of an aural pressure modulatingagent, or pharmaceutically acceptable prodrug or salt thereof, by weightof the composition. In some embodiments, a pharmaceutical composition ordevice described above comprises about 50% of an aural pressuremodulating agent, or pharmaceutically acceptable prodrug or saltthereof, by weight of the composition.

In some embodiments, a pharmaceutical composition or device describedabove has a pH between about 5.5 to about 8.0. In some embodiments, apharmaceutical composition or device described above has a pH betweenabout 6.0 to about 8.0. In some embodiments, a pharmaceuticalcomposition or device described above has a pH between about 6.0 toabout 7.6. In some embodiments, a pharmaceutical composition or devicedescribed above has a pH between about 7.0 to about 7.6.

In some embodiments, a pharmaceutical composition or device describedabove contains less than 100 colony forming units (cfu) ofmicrobiological agents per gram of formulation. In some embodiments, apharmaceutical composition or device described above contains less than50 colony forming units (cfu) of microbiological agents per gram offormulation. In some embodiments, a pharmaceutical composition or devicedescribed above contains less than 10 colony forming units (cfu) ofmicrobiological agents per gram of formulation.

In some embodiments, a pharmaceutical composition or device describedabove contains less than 5 endotoxin units (EU) per kg of body weight ofa subject. In some embodiments, a pharmaceutical composition or devicedescribed above contains less than 4 endotoxin units (EU) per kg of bodyweight of a subject.

In some embodiments a pharmaceutical composition or device describedabove provides a gelation temperature between about between about 19° C.to about 42° C. In some embodiments a pharmaceutical composition ordevice described above provides a gelation temperature between aboutbetween about 19° C. to about 37° C. In some embodiments apharmaceutical composition or device described above provides a gelationtemperature between about between about 19° C. to about 30° C.

In some embodiments, the pharmaceutical composition or device is anauris-acceptable thermoreversible gel. In some embodiments, thepolyoxyethylene-polyoxypropylene triblock copolymer is biodegradableand/or bioeliminated (e.g., the copolymer is eliminated from the body bya biodegradation process, e.g., elimination in the urine, the feces orthe like). In some embodiments, a pharmaceutical composition or devicedescribed herein further comprises a mucoadhesive. In some embodiments,a pharmaceutical composition or device described herein furthercomprises a penetration enhancer. In some embodiments, a pharmaceuticalcomposition or device described herein further comprises a thickeningagent. In some embodiments, a pharmaceutical composition or devicedescribed herein further comprises a dye.

In some embodiments, a pharmaceutical composition or device describedherein further comprises a drug delivery device selected from a needleand syringe, a pump, a microinjection device, a wick, an in situ formingspongy material or combinations thereof.

In some embodiments, a pharmaceutical composition or device describedherein is a pharmaceutical composition or device wherein the auralpressure modulating agent, or pharmaceutically acceptable salt thereof,has limited or no systemic release, systemic toxicity, poor PKcharacteristics, or combinations thereof. In some embodiments of thepharmaceutical compositions or devices described herein, the auralpressure modulating agent is in the form of a neutral molecule, freebase, a free acid, a salt, a prodrug, or a combination thereof. In someembodiments of the pharmaceutical compositions or devices describedherein, the aural pressure modulating agent is administered in the formof a phosphate or ester prodrug. In some embodiments pharmaceuticalcompositions or devices described herein comprise one or more auralpressure modulating agent, or pharmaceutically acceptable salt thereof,prodrug or combination thereof as an immediate release agent.

In some embodiments, pharmaceutical compositions or devices describedherein further comprise an additional therapeutic agent. In someembodiments, the additional therapeutic agent is a diuretic, ananti-emetic agent, an anti-vertigo agent, a local acting anestheticagent, an anti-anxiety agent, a corticosteroid, an anti-histamine, orcombinations thereof. In some embodiments, the additional therapeuticagent is an otoprotectant, a Na/K ATPase modulator, a KCNQ channelmodulator, or combinations thereof.

In some embodiments, pharmaceutical compositions or devices describedherein are pharmaceutical compositions or devices wherein the pH of thepharmaceutical composition or device is between about 6.0 to about 7.6.

In some embodiments of the pharmaceutical compositions or devicesdescribed herein, the ratio of a polyoxyethylene-polyoxypropylenetriblock copolymer of general formula E 106 P70 E106 to a thickeningagent is from about 40:1 to about 5:1. In some embodiments, thethickening agent is carboxymethyl cellulose, hydroxypropyl cellulose orhydroxypropyl methylcellulose.

In some embodiments, the otic disease or condition is Meniere's disease,sudden sensorineural hearing loss, noise induced hearing loss, agerelated hearing loss, auto immune ear disease or tinnitus.

Also provided herein is a method of treating an otic disease orcondition associated with an imbalance in aural pressure comprisingadministering to an individual in need thereof an intratympaniccomposition or device comprising a therapeutically effective amount ofaural pressure modulating agent, the composition or device comprisingsubstantially low degradation products of aural pressure modulatingagent, the composition or device further comprising two or morecharacteristics selected from:

-   -   (i) between about 0.1% to about 10% by weight of the aural        pressure modulating agent, or pharmaceutically acceptable        prodrug or salt thereof;    -   (ii) between about 14% to about 21% by weight of a        polyoxyethylene-polyoxypropylene triblock copolymer of general        formula E106 P70 E106;    -   (iii) sterile water, q.s., buffered to provide a pH between        about 5.5 and about 8.0;    -   (iv) multiparticulate aural pressure modulating agent;    -   (v) a gelation temperature between about 19° C. to about 42° C.;    -   (vi) less than about 50 colony forming units (cfu) of        microbiological agents per gram of formulation, and    -   (vii) less than about 5 endotoxin units (EU) per kg of body        weight of a subject.

In some embodiments of the methods described herein, the aural pressuremodulating agent is released from the composition or device for a periodof at least 3 days. In some embodiments of the methods described herein,the aural pressure modulating agent is released from the composition ordevice for a period of at least 5 days. In some embodiments of themethods described herein, the aural pressure modulating agent isreleased from the composition or device for a period of at least 10days. In some embodiments of the method described above, the auralpressure modulating agent is essentially in the form of micronizedparticles.

In some embodiments of the methods described herein, the composition isadministered across the round window. In some embodiments of the methodsdescribed herein, the otic disease or condition is Meniere's disease,sudden sensorineural hearing loss, noise induced hearing loss, agerelated hearing loss, auto immune ear disease or tinnitus.

BRIEF DESCRIPTION OF FIGURES

FIG. 1. illustrates a comparison of non-sustained release and sustainedrelease formulations.

FIG. 2 illustrates the effect of concentration on the viscosity ofaqueous solutions of Blanose refined CMC.

FIG. 3 illustrates the effect of concentration on the viscosity ofaqueous solutions of Methocel.

FIG. 4 illustrates the anatomy of the ear

DETAILED DESCRIPTION OF THE INVENTION

Provided herein are controlled release aural pressure modulatingcompositions and formulations to treat otic diseases or conditionsassociated with an imbalance in aural pressure (e.g., fluid homeostasisdisorders of the ear), including Meniere's Disease, endolymphatichydrops, progressive hearing loss, dizziness, vertigo, tinnitus andsimilar conditions. In some instances, an imbalance in aural pressure inthe ear is a result of an imbalance in fluid homeostasis control in theinner ear, or auris interna. In some instances, an imbalance in auralpressure is a result of an imbalance in fluid homeostasis control in themiddle ear, or auris media. In some instances, an imbalance in auralpressure in the ear causes a feeling of fullness in the ear. In someinstances, an imbalance in aural pressure in the ear causes nauseaand/or vertigo and/or tinnitus.

Systemic routes via oral, intravenous or intramuscular routes arecurrently used to deliver aural pressure modulating therapeutic agents.Systemic drug administration may create a potential inequality in drugconcentration with higher circulating levels in the serum, and lowerlevels in the target auris interna organ structures. As a result, fairlylarge amounts of drug are required to overcome this inequality in orderto deliver sufficient, therapeutically effective quantities to the innerear. In addition, systemic drug administration may increase thelikelihood of systemic toxicities and adverse side effects as a resultof the high serum amounts required to effectuate sufficient localdelivery to the target site. Systemic toxicities may also occur as aresult of liver breakdown and processing of the therapeutic agents,forming toxic metabolites that effectively erase any benefit attainedfrom the administered therapeutic. Systemic administration of auralpressure modulators may result in the unwanted modulation of other organsystems where water homeostasis is vital, such as the cardiovascular orrenal system. For example, administration of the vasopressin antagonistconivaptan is indicated in patients with hypervolemic hyponatremiaarising from an underlying cause, such as congestive heart failure.Systemic administration of a vasopressin antagonist to a patientsuffering only from fluid homeostasis disorders of the ear could resultin hypovolemia, hypematremia or hypotension in the patient, due tosystemic effects of vasopressin antagonists on the renal system.

To overcome the toxic and attendant side effects of systemic delivery,disclosed herein are methods and compositions for local delivery ofaural pressure modulators to auris interna structures. Access to, forexample, the vestibular and cochlear apparatus occurs through the aurismedia, including the oval window/stapes footplate, the annular ligamentand through the otic capsule/temporal bone, and the round windowmembrane of the auris interna.

In addition, localized treatment of the auris interna also affords theuse of previously undesired therapeutic agents, including agents withpoor pK profiles, poor uptake, low systemic release and/or toxicityissues. Because of the localized targeting of the aural pressuremodulating formulations and compositions, as well as the biologicalblood barrier present in the auris interna, the risk of adverse effectswill be reduced as a result of treatment with previously characterizedtoxic or ineffective aural pressure modulators. Accordingly, alsocontemplated within the scope of the embodiments herein is the use ofaural pressure modulating agents in the treatment of fluid homeostasisotic disorders that have been previously rejected by practitionersbecause of adverse effects or ineffectiveness of the aural pressuremodulators (e.g., vasopressin receptor modulators).

Also included within the embodiments disclosed herein is the use ofadditional auris interna agents in combination with the aural pressuremodulating formulations and compositions disclosed herein. When used,such agents assist in the treatment of hearing or equilibrium loss ordysfunction as a result of a fluid homeostasis disorder, includingvertigo, tinnitus, hearing loss, balance disorders or combinationsthereof. Accordingly, agents that ameliorate or reduce the effects ofvertigo, tinnitus, hearing loss, balance disorders, or combinationsthereof are also contemplated to be used in combination with the auralpressure modulators, including steroids, anti-emetic agents,corticosteroids, local acting anesthetic agents, chemotherapeuticagents, including cytoxan, azathiaprine or methotrexate; andcombinations thereof.

In addition, the aural pressure modulating pharmaceutical compositionsor formulations included herein also include carriers, adjuvants, suchas preserving, stabilizing, wetting or emulsifying agents, solutionpromoters, salts for regulating the osmotic pressure, and/or buffers.Such carriers, adjuvants, and other excipients will be compatible withthe environment in the auris interna. Accordingly, specificallycontemplated are carriers, adjuvants and excipients that lackototoxicity or are minimally ototoxic in order to allow effectivetreatment of the otic disorders contemplated herein with minimal sideeffects in the targeted regions or areas. To prevent ototoxicity, auralpressure modulating pharmaceutical compositions or formulationsdisclosed herein are optionally targeted to distinct regions of theauris interna, including but not limited to the vestibular bony andmembranous labyrinths, cochlear bony and membranous labyrinths and otheranatomical or physiological structures located within the auris interna.

In some instances, systemic drug administration creates a potentialinequality in drug concentration with higher circulating levels in theserum, and lower levels in the target auris media and auris internaorgan structures. As a result, fairly large amounts of drug are requiredto overcome this inequality in order to deliver sufficient,therapeutically effective quantities to the inner ear. In addition,systemic drug administration may increase the likelihood of systemictoxicities and adverse side effects as a result of the high serumamounts required to effectuate sufficient local delivery to the targetsite. Systemic toxicities may also occur as a result of liver breakdownand processing of the therapeutic agents, forming toxic metabolites thateffectively erase any benefit attained from the administeredtherapeutic.

To overcome the toxic and attendant side effects of systemic delivery,disclosed herein are methods and compositions and devices for localdelivery of therapeutic agents to targeted auris structures. Access to,for example, the vestibular and cochlear apparatus will occur throughthe auris media including round window membrane, the oval window/stapesfootplate, the annular ligament and through the otic capsule/temporalbone.

Intratympanic injection of therapeutic agents is the technique ofinjecting a therapeutic agent behind the tympanic membrane into theauris media and/or auris interna. Despite early success with thistechnique (Schuknecht, Laryngoscope (1956) 66, 859-870) some challengesdo remain. For example, access to the round window membrane, the site ofdrug absorption into the auris interna, can be challenging.

However, intra-tympanic injections create several unrecognized problemsnot addressed by currently available treatment regimens, such aschanging the osmolarity and pH of the perilymph and endolymph, andintroducing pathogens and endotoxins that directly or indirectly damageinner ear structures. One of the reasons the art may not have recognizedthese problems is that there are no approved intra-tympaniccompositions: the inner ear provides sui generis formulation challenges.Thus, compositions developed for other parts of the body have little tono relevance for an intra-tympanic composition.

There is no guidance in the prior art regarding requirements (e.g.,level of sterility, pH, osmolarity) for otic formulations that aresuitable for administration to humans. There is wide anatomicaldisparity between the ears of animals across species. A consequence ofthe inter-species differences in auditory structures is that animalmodels of inner ear disease are often unreliable as a tool for testingtherapeutics that are being developed for clinical approval.

Provided herein are otic formulations that meet stringent criteria forpH, osmolarity, ionic balance, sterility, endotoxin and/or pyrogenlevels. The auris compositions described herein are compatible with themicroenvironment of the inner ear (e.g., the perilymph) and are suitablefor administration to humans. In some embodiments, the formulationsdescribed herein comprise dyes and aid visualization of the administeredcompositions obviating the need for invasive procedures (e.g., removalof perilymph) during preclinical and/or clinical development ofintratympanic therapeutics.

Provided herein are controlled release aural pressure modulating agentformulations and compositions to locally treat targeted aurisstructures, thereby avoiding side effects as a result of systemicadministration of the aural pressure modulating agent formulations andcompositions. The locally applied aural pressure modulating agentformulations and compositions and devices are compatible with thetargeted auris structures, and administered either directly to thedesired targeted auris structure, e.g. the cochlear region, the tympaniccavity or the external ear, or administered to a structure in directcommunication with areas of the auris interna, including but not limitedto the round window membrane, the crista fenestrae cochleae or the ovalwindow membrane. By specifically targeting an auris structure, adverseside effects as a result of systemic treatment are avoided. Moreover,clinical studies have shown the benefit of having long term exposure ofdrug to the perilymph of the cochlea, for example with improved clinicalefficacy of sudden hearing loss when the therapeutic agent is given onmultiple occasions. Thus, by providing a controlled release auralpressure modulating agent formulation or composition to treat oticdisorders, a constant, variable and/or extended source of aural pressuremodulating agent is provided to the individual or patient suffering froman otic disorder, reducing or eliminating variabilities in treatment.Accordingly, one embodiment disclosed herein is to provide a compositionthat enables at least one aural pressure modulating agent to be releasedin therapeutically effective doses either at variable or constant ratessuch as to ensure a continuous release of the at least one agent. Insome embodiments, the aural pressure modulating agents disclosed hereinare administered as an immediate release formulation or composition. Inother embodiments, the aural pressure modulating agents are administeredas a sustained release formulation, released either continuously,variably or in a pulsatile manner, or variants thereof. In still otherembodiments, aural pressure modulating agent formulation is administeredas both an immediate release and sustained release formulation, releasedeither continuously, variably or in a pulsatile manner, or variantsthereof. The release is optionally dependent on environmental orphysiological conditions, for example, the external ionic environment(see, e.g. Oros® release system, Johnson & Johnson).

In addition, localized treatment of the targeted auris structure alsoaffords the use of previously undesired therapeutic agents, includingagents with poor pK profiles, poor uptake, low systemic release and/ortoxicity issues. Because of the localized targeting of the auralpressure modulating agent formulations and compositions and devices, aswell as the biological blood barrier present in the auris interna, therisk of adverse effects will be reduced as a result of treatment withpreviously characterized toxic or ineffective aural pressure modulatingagents. Accordingly, also contemplated within the scope of theembodiments herein is the use of aural pressure modulating agents in thetreatment of homeostasis-related otic disorders that have beenpreviously rejected by practitioners because of adverse effects orineffectiveness of the aural pressure modulating agent.

Also included within the embodiments disclosed herein is the use ofadditional auris-compatible agents in combination with the auralpressure modulating agent formulations and compositions and devicesdisclosed herein. When used, such agents assist in the treatment ofhearing or equilibrium loss or dysfunction as a result of an autoimmunedisorder, including vertigo, tinnitus, hearing loss, balance disorders,infections, or combinations thereof. Accordingly, agents that ameliorateor reduce the effects of vertigo, tinnitus, hearing loss, balancedisorders, infections, inflammatory response or combinations thereof arealso contemplated to be used in combination with the aural pressuremodulating agent(s), including a diuretic, an anti-emetic agent, ananti-vertigo agent, a local acting anesthetic agent, an anti-anxietyagent, a corticosteroid, an anti-histamine an otoprotectant, a Na/KATPase modulator, a KCNQ channel modulator, or combinations thereof.

In addition, the auris-acceptable controlled-release aural pressuremodulating agent formulations and treatments described herein areprovided to the target ear region of the individual in need, includingthe inner ear, and the individual in need is additionally administeredan oral dose of aural pressure modulating agent. In some embodiments,the oral dose of aural pressure modulating agent is administered priorto administration of the auris-acceptable controlled-release auralpressure modulating agent formulation, and then the oral dose is taperedoff over the period of time that the auris-acceptable controlled-releaseaural pressure modulating agent formulation is provided. Alternatively,the oral dose of aural pressure modulating agent is administered duringadministration of the auris-acceptable controlled-release aural pressuremodulating agent formulation, and then the oral dose is tapered off overthe period of time that the auris-acceptable controlled-release auralpressure modulating agent formulation is provided. Alternatively, theoral dose of aural pressure modulating agent is administered afteradministration of the auris-acceptable controlled-release aural pressuremodulating agent formulation has been initiated, and then the oral doseis tapered off over the period of time that the auris-acceptablecontrolled-release aural pressure modulating agent formulation isprovided.

In addition, the aural pressure modulating agent pharmaceuticalcompositions or formulations or devices included herein also includecarriers, adjuvants, such as preserving, stabilizing, wetting oremulsifying agents, solution promoters, salts for regulating the osmoticpressure, and/or buffers. Such carriers, adjuvants, and other excipientswill be compatible with the environment in the targeted aurisstructure(s). Accordingly, specifically contemplated are carriers,adjuvants and excipients that lack ototoxicity or are minimally ototoxicin order to allow effective treatment of the otic disorders contemplatedherein with minimal side effects in the targeted regions or areas. Toprevent ototoxicity, aural pressure modulating agent pharmaceuticalcompositions or formulations or devices disclosed herein are optionallytargeted to distinct regions of the targeted auris structures, includingbut not limited to the tympanic cavity, vestibular bony and membranouslabyrinths, cochlear bony and membranous labyrinths and other anatomicalor physiological structures located within the auris interna.

Certain Definitions

The term “auris-acceptable” with respect to a formulation, compositionor ingredient, as used herein, includes having no persistent detrimentaleffect on the auris interna (or inner ear) of the subject being treated.By “auris-pharmaceutically acceptable,” as used herein, refers to amaterial, such as a carrier or diluent, which does not abrogate thebiological activity or properties of the compound in reference to theauris interna (or inner ear), and is relatively or is reduced intoxicity to the auris interna (or inner ear), i.e., the material isadministered to an individual without causing undesirable biologicaleffects or interacting in a deleterious manner with any of thecomponents of the composition in which it is contained.

As used herein, amelioration or lessening of the symptoms of aparticular otic disease, disorder or condition by administration of aparticular compound or pharmaceutical composition refers to any decreaseof severity, delay in onset, slowing of progression, or shortening ofduration, whether permanent or temporary, lasting or transient that isattributed to or associated with administration of the compound orcomposition.

As used herein, the terms “aural pressure modulating agent” or “auralpressure modulator” are used as synonyms and do not define the degree ofefficacy. The aural pressure modulator also includes compounds thatmodulate the expression or post-transcriptional processing of a fluidhomeostasis protein, including vasopressin and estrogen-related receptorbeta protein. Additionally, vasopressin receptor or estrogen-relatedreceptor beta modulators include compounds that influence vasopressinreceptor or estrogen-related receptor beta signalling or downstreamfunctions under the control of the vasopressin receptor orestrogen-related receptor beta, such as aquaporin function. Vasopressinreceptor or estrogen-related receptor beta modulating agents includescompounds that increase and/or decrease vasopressin receptor orestrogen-related receptor beta function, including antagonists,inhibitors, agonists, partial agonists and the like.

“Antioxidants” are auris-pharmaceutically acceptable antioxidants, andinclude, for example, butylated hydroxytoluene (BHT), sodium ascorbate,ascorbic acid, sodium metabisulfite and tocopherol. In certainembodiments, antioxidants enhance chemical stability where required.Antioxidants are also used to counteract the ototoxic effects of certaintherapeutic agents, including agents that are used in combination withthe aural pressure modulating agents disclosed herein.

“Auris interna” refers to the inner ear, including the cochlea and thevestibular labyrinth, and the round window that connects the cochleawith the middle ear.

“Auris-interna bioavailability” refers to the percentage of theadministered dose of compounds disclosed herein that becomes availablein the inner ear of the animal or human being studied.

“Auris media” refers to the middle ear, including the tympanic cavity,auditory ossicles and oval window, which connects the middle ear withthe inner ear.

“Blood plasma concentration” refers to the concentration of compoundsprovided herein in the plasma component of blood of a subject.

“Carrier materials” are excipients that are compatible with the auralpressure modulator, the auris interna and the release profile propertiesof the auris-acceptable pharmaceutical formulations. Such carriermaterials include, e.g., binders, suspending agents, disintegrationagents, filling agents, surfactants, solubilizers, stabilizers,lubricants, wetting agents, diluents, and the like.“Auris-pharmaceutically compatible carrier materials” include, but arenot limited to, acacia, gelatin, colloidal silicon dioxide, calciumglycerophosphate, calcium lactate, maltodextrin, glycerine, magnesiumsilicate, polyvinylpyrrolidone (PVP), cholesterol, cholesterol esters,sodium caseinate, soy lecithin, taurocholic acid, phosphatidylcholine,sodium chloride, tricalcium phosphate, dipotassium phosphate, celluloseand cellulose conjugates, sugars sodium stearoyl lactylate, carrageenan,monoglyceride, diglyceride, pregelatinized starch, and the like.

The term “diluent” refers to chemical compounds that are used to dilutethe aural pressure modulator prior to delivery and which are compatiblewith the auris interna.

“Dispersing agents,” and/or “viscosity modulating agents” are materialsthat control the diffusion and homogeneity of the aural pressuremodulator through liquid media. Examples of diffusionfacilitators/dispersing agents include but are not limited tohydrophilic polymers, electrolytes, Tween® 60 or 80, PEG,polyvinylpyrrolidone (PVP; commercially known as Plasdone®), and thecarbohydrate-based dispersing agents such as, for example, hydroxypropylcelluloses (e.g., HPC, HPC-SL, and HPC-L), hydroxypropylmethylcelluloses (e.g., HPMC K100, HPMC K4M, HPMC K15M, and HPMC K100M),carboxymethylcellulose sodium, methylcellulose, hydroxyethylcellulose,hydroxypropylcellulose, hydroxypropylmethylcellulose phthalate,hydroxypropylmethylcellulose acetate stearate (HPMCAS), noncrystallinecellulose, magnesium aluminum silicate, triethanolamine, polyvinylalcohol (PVA), vinyl pyrrolidone/vinyl acetate copolymer (S630),4-(1,1,3,3-tetramethylbutyl)-phenol polymer with ethylene oxide andformaldehyde (also known as tyloxapol), poloxamers (e.g., PluronicsF68®, F88®, and F108®, which are block copolymers of ethylene oxide andpropylene oxide); and poloxamines (e.g., Tetronic 908®, also known asPoloxamine 908®, which is a tetrafunctional block copolymer derived fromsequential addition of propylene oxide and ethylene oxide toethylenediamine (BASF Corporation, Parsippany, N.J.)),polyvinylpyrrolidone K12, polyvinylpyrrolidone K17, polyvinylpyrrolidoneK25, or polyvinylpyrrolidone K30, polyvinylpyrrolidone/vinyl acetatecopolymer (S-630), polyethylene glycol, e.g., the polyethylene glycolhas a molecular weight of about 300 to about 6000, or about 3350 toabout 4000, or about 7000 to about 5400, sodium carboxymethylcellulose,methylcellulose, polysorbate-80, sodium alginate, gums, such as, e.g.,gum tragacanth and gum acacia, guar gum, xanthans, including xanthangum, sugars, cellulosics, such as, sodium carboxymethylcellulose,methylcellulose, sodium carboxymethylcellulose, polysorbate-80, sodiumalginate, polyethoxylated sorbitan monolaurate, polyethoxylated sorbitanmonolaurate, povidone, carbomers, polyvinyl alcohol (PVA), alginates,chitosans and combinations thereof. Plasticizers such as cellulose ortriethyl cellulose are also be used as dispersing agents. Dispersingagents useful in liposomal dispersions and self-emulsifying dispersionsof the aural pressure modulators disclosed herein are dimyristoylphosphatidyl choline, natural phosphatidyl choline from eggs, naturalphosphatidyl glycerol from eggs, cholesterol and isopropyl myristate.

“Drug absorption” or “absorption” refers to the process of movement ofthe aural pressure modulating agents from the localized site ofadministration, by way of example only, the round window membrane of theinner ear, and across a barrier (the round window membranes, asdescribed below) into the auris interna or inner ear structures. Theterms “co-administration” or the like, as used herein, are meant toencompass administration of the aural pressure modulating agents to asingle patient, and are intended to include treatment regimens in whichthe aural pressure modulating agents are administered by the same ordifferent route of administration or at the same or different time.

The terms “effective amount” or “therapeutically effective amount,” asused herein, refer to a sufficient amount of the aural pressuremodulating agent being administered that would be expected to relieve tosome extent one or more of the symptoms of the disease or conditionbeing treated. For example, the result of administration of the auralpressure modulating agents disclosed herein is reduction and/oralleviation of the signs, symptoms, or causes of improper fluidhomeostasis in the inner ear. For example, an “effective amount” fortherapeutic use is the amount of the aural pressure modulator, includinga formulation as disclosed herein required to provide a decrease oramelioration in disease symptoms without undue adverse side effects. Theterm “therapeutically effective amount” includes, for example, aprophylactically effective amount. An “effective amount” of an auralpressure modulator composition disclosed herein is an amount effectiveto achieve a desired pharmacologic effect or therapeutic improvementwithout undue adverse side effects. It is understood that “an effectiveamount” or “a therapeutically effective amount” varies, in someembodiments, from subject to subject, due to variation in metabolism ofthe compound administered, age, weight, general condition of thesubject, the condition being treated, the severity of the conditionbeing treated, and the judgment of the prescribing physician. It is alsounderstood that “an effective amount” in an extended release dosingformat may differ from “an effective amount” in an immediate releasedosing format based upon pharmacokinetic and pharmacodynamicconsiderations.

The terms “enhance” or “enhancing” refers to an increase or prolongationof either the potency or duration of a desired effect of the auralpressure modulators or a diminution of any adverse symptomatology thatis consequent upon the administration of the aural pressure modulatingagent. Thus, in regard to enhancing the effect of the aural pressuremodulators disclosed herein, the term “enhancing” refers to the abilityto increase or prolong, either in potency or duration, the effect ofother therapeutic agents that are used in combination with the auralpressure modulators disclosed herein. An “enhancing-effective amount,”as used herein, refers to an amount of aural pressure modulator or othertherapeutic agent that is adequate to enhance the effect of anothertherapeutic agent or aural pressure modulator in a desired system. Whenused in a patient, amounts effective for this use will depend on theseverity and course of the disease, disorder or condition, previoustherapy, the patient's health status and response to the drugs, and thejudgment of the treating physician.

The term “inhibiting” includes preventing, slowing, or reversing thedevelopment of a condition, for example, improper inner ear fluidhomeostasis, or advancement of a condition in a patient necessitatingtreatment.

The terms “kit” and “article of manufacture” are used as synonyms.

The term “modulate” includes the interaction of a ligand with areceptor, the fluid homeostasis, or other direct or indirect targetsthat alter the activity of the fluid homeostasis, including, by way ofexample only, to inhibit the activity of vasopressin peptide, to inhibitthe expression or post-transcriptional processing of the vasopressionreceptor protein. Additionally, modulation includes influencing thesignalling or downstream functions under the control of, for example,vasopressin receptor or estrogen-related receptor beta, such asaquaporin function.

“Pharmacodynamics” refers to the factors which determine the biologicresponse observed relative to the concentration of drug at the desiredsite within the auris media and/or auris interna.

“Pharmacokinetics” refers to the factors which determine the attainmentand maintenance of the appropriate concentration of drug at the desiredsite within the auris media and/or auris interna.

In prophylactic applications, compositions containing the aural pressuremodulator described herein are administered to a patient susceptible toor otherwise at risk of a particular disease, disorder or condition, forexample, a disorder of inner ear fluid homeostasis, or patients that aresuffering from a disease or symptoms known to be characteristic of adisorder of inner ear fluid homeostasis, including by way of exampleonly, Meniere's Disease, endolymphatic hydrops, progressive hearingloss, dizziness, vertigo, and tinnitus. Such an amount is defined to bea “prophylactically effective amount or dose.” In this use, the preciseamounts also depend on the patient's state of health, weight, and thelike.

As used herein, a “pharmaceutical device” includes any compositiondescribed herein that, upon administration to an ear, provides areservoir for extended release of an active agent described herein.

A “prodrug” refers to an aural pressure modulating agent that isconverted into the parent drug in vivo. In certain embodiments, aprodrug is enzymatically metabolized by one or more steps or processesto the biologically, pharmaceutically or therapeutically active form ofthe compound. To produce a prodrug, a pharmaceutically active compoundis modified such that the active compound will be regenerated upon invivo administration. In one embodiment, the prodrug is designed to alterthe metabolic stability or the transport characteristics of a drug, tomask side effects or toxicity, or to alter other characteristics orproperties of a drug. Compounds provided herein, in some embodiments,are derivatized into suitable prodrugs.

“Solubilizers” refer to auris-acceptable compounds such as triacetin,triethylcitrate, ethyl oleate, ethyl caprylate, sodium lauryl sulfate,sodium doccusate, vitamin E TPGS, dimethylacetamide,N-methylpyrrolidone, N-hydroxyethylpyrrolidone, polyvinylpyrrolidone,hydroxypropylmethyl cellulose, hydroxypropyl cyclodextrins, ethanol,n-butanol, isopropyl alcohol, cholesterol, bile salts, polyethyleneglycol 200-600, glycofurol, transcutol, propylene glycol, and dimethylisosorbide and the like that assist or increase the solubility of theaural pressure modulators disclosed herein.

“Stabilizers” refers to compounds such as any antioxidation agents,buffers, acids, preservatives and the like that are compatible with theenvironment of the auris interna. Stabilizers include but are notlimited to agents that will do any of (1) improve the compatibility ofexcipients with a container, or a delivery system, including a syringeor a glass bottle, (2) improve the stability of a component of thecomposition, or (3) improve formulation stability.

“Steady state,” as used herein, is when the amount of drug administeredto the auris interna is equal to the amount of drug eliminated withinone dosing interval resulting in a plateau or constant levels of drugexposure within the targeted structure.

As used herein, the term “subject” is used to mean an animal, preferablya mammal, including a human or non-human. The terms patient and subjectmay be used interchangeably.

“Surfactants” refer to compounds that are auris-acceptable, such assodium lauryl sulfate, sodium docusate, Tween 60 or 80, triacetin,vitamin E TPGS, sorbitan monooleate, polyoxyethylene sorbitanmonooleate, polysorbates, polaxomers, bile salts, glyceryl monostearate,copolymers of ethylene oxide and propylene oxide, e.g., Pluronic®(BASF), and the like. Some other surfactants include polyoxyethylenefatty acid glycerides and vegetable oils, e.g., polyoxyethylene (60)hydrogenated castor oil; and polyoxyethylene alkylethers and alkylphenylethers, e.g., octoxynol 10, octoxynol 40. In some embodiments,surfactants are included to enhance physical stability or for otherpurposes.

The terms “treat,” “treating” or “treatment,” as used herein, includealleviating, abating or ameliorating a disease or condition, for examplea disorder of inner ear fluid homeostasis, symptoms, preventingadditional symptoms, ameliorating or preventing the underlying metaboliccauses of symptoms, inhibiting the disease or condition, e.g., arrestingthe development of the disease or condition, relieving the disease orcondition, causing regression of the disease or condition, relieving acondition caused by the disease or condition, or stopping the symptomsof the disease or condition either prophylactically and/ortherapeutically.

Other objects, features, and advantages of the methods and compositionsdescribed herein will become apparent from the following detaileddescription. It should be understood, however, that the detaileddescription and the specific examples, while indicating specificembodiments, are given by way of illustration only.

Anatomy of the Ear

As shown FIG. 4, the outer ear is the external portion of the organ andis composed of the pinna (auricle), the auditory canal (externalauditory meatus) and the outward facing portion of the tympanicmembrane, also known as the ear drum. The pinna, which is the fleshypart of the external ear that is visible on the side of the head,collects sound waves and directs them toward the auditory canal. Thus,the function of the outer ear, in part, is to collect and direct soundwaves towards the tympanic membrane and the middle ear.

The middle ear is an air-filled cavity, called the tympanic cavity,behind the tympanic membrane. The tympanic membrane, also known as theear drum, is a thin membrane that separates the external ear from themiddle ear. The middle ear lies within the temporal bone, and includeswithin this space the three ear bones (auditory ossicles): the malleus,the incus and the stapes. The auditory ossicles are linked together viatiny ligaments, which form a bridge across the space of the tympaniccavity. The malleus, which is attached to the tympanic membrane at oneend, is linked to the incus at its anterior end, which in turn is linkedto the stapes. The stapes is attached to the oval window, one of twowindows located within the tympanic cavity. A fibrous tissue layer,known as the annular ligament connects the stapes to the oval window.Sound waves from the outer ear first cause the tympanic membrane tovibrate. The vibration is transmitted across to the cochlea through theauditory ossicles and oval window, which transfers the motion to thefluids in the auris interna. Thus, the auditory ossicles are arranged toprovide a mechanical linkage between the tympanic membrane and the ovalwindow of the fluid-filled auris interna, where sound is transformed andtransduced to the auris interna for further processing. Stiffness,rigidity or loss of movement of the auditory ossicles, tympanic membraneor oval window leads to hearing loss, e.g. otosclerosis, or rigidity ofthe stapes bone.

The tympanic cavity also connects to the throat via the eustachian tube.The eustachian tube provides the ability to equalize the pressurebetween the outside air and the middle ear cavity. The round window, acomponent of the auris interna but which is also accessible within thetympanic cavity, opens into the cochlea of the auris interna. The roundwindow is covered by round window membrane, which consists of threelayers: an external or mucous layer, an intermediate or fibrous layer,and an internal membrane, which communicates directly with the cochlearfluid. The round window, therefore, has direct communication with theauris interna via the internal membrane.

Movements in the oval and round window are interconnected, i.e. as thestapes bone transmits movement from the tympanic membrane to the ovalwindow to move inward against the auris interna fluid, the round window(round window membrane) is correspondingly pushed out and away from thecochlear fluid. This movement of the round window allows movement offluid within the cochlea, which leads in turn to movement of thecochlear inner hair cells, allowing hearing signals to be transduced.Stiffness and rigidity in round window membrane leads to hearing lossbecause of the lack of ability of movement in the cochlear fluid. Recentstudies have focused on implanting mechanical transducers onto the roundwindow, which bypasses the normal conductive pathway through the ovalwindow and provides amplified input into the cochlear chamber.

Auditory signal transduction takes place in the auris interna. Thefluid-filled auris interna, or inner ear, consists of two majorcomponents: the cochlear and the vestibular apparatus. The auris internais located in part within the osseous or bony labyrinth, an intricateseries of passages in the temporal bone of the skull. The vestibularapparatus is the organ of balance and consists of the threesemi-circular canals and the vestibule. The three semi-circular canalsare arranged relative to each other such that movement of the head alongthe three orthogonal planes in space can be detected by the movement ofthe fluid and subsequent signal processing by the sensory organs of thesemi-circular canals, called the crista ampullaris. The cristaampullaris contains hair cells and supporting cells, and is covered by adome-shaped gelatinous mass called the cupula. The hairs of the haircells are embedded in the cupula. The semi-circular canals detectdynamic equilibrium, the equilibrium of rotational or angular movements.

When the head turns rapidly, the semicircular canals move with the head,but endolymph fluid located in the membranous semi-circular canals tendsto remain stationary. The endolymph fluid pushes against the cupula,which tilts to one side. As the cupula tilts, it bends some of the hairson the hair cells of the crista ampullaris, which triggers a sensoryimpulse. Because each semicircular canal is located in a differentplane, the corresponding crista ampullaris of each semi-circular canalresponds differently to the same movement of the head. This creates amosaic of impulses that are transmitted to the central nervous system onthe vestibular branch of the vestibulocochlear nerve. The centralnervous system interprets this information and initiates the appropriateresponses to maintain balance. Of importance in the central nervoussystem is the cerebellum, which mediates the sense of balance andequilibrium.

The vestibule is the central portion of the auris interna and containsmechanoreceptors bearing hair cells that ascertain static equilibrium,or the position of the head relative to gravity. Static equilibriumplays a role when the head is motionless or moving in a straight line.The membranous labyrinth in the vestibule is divided into two sac-likestructures, the utricle and the saccule. Each structure in turn containsa small structure called a macula, which is responsible for maintenanceof static equilibrium. The macula consists of sensory hair cells, whichare embedded in a gelatinous mass (similar to the cupula) that coversthe macula. Grains of calcium carbonate, called otoliths, are embeddedon the surface of the gelatinous layer.

When the head is in an upright position, the hairs are straight alongthe macula. When the head tilts, the gelatinous mass and otoliths tiltscorrespondingly, bending some of the hairs on the hair cells of themacula. This bending action initiates a signal impulse to the centralnervous system, which travels via the vestibular branch of thevestibulocochlear nerve, which in turn relays motor impulses to theappropriate muscles to maintain balance.

The cochlea is the portion of the auris interna related to hearing. Thecochlea is a tapered tube-like structure which is coiled into a shaperesembling a snail. The inside of the cochlea is divided into threeregions, which is further defined by the position of the vestibularmembrane and the basilar membrane. The portion above the vestibularmembrane is the scala vestibuli, which extends from the oval window tothe apex of the cochlea and contains perilymph fluid, an aqueous liquidlow in potassium and high in sodium content. The basilar membranedefines the scala tympani region, which extends from the apex of thecochlea to the round window and also contains perilymph. The basilarmembrane contains thousands of stiff fibers, which gradually increase inlength from the round window to the apex of the cochlea. The fibers ofthe basement membrane vibrate when activated by sound. In between thescala vestibuli and the scala tympani is the cochlear duct, which endsas a closed sac at the apex of the cochlea. The cochlear duct containsendolymph fluid, which is similar to cerebrospinal fluid and is high inpotassium.

The organ of Corti, the sensory organ for hearing, is located on thebasilar membrane and extends upward into the cochlear duct. The organ ofCorti contains hair cells, which have hairlike projections that extendfrom their free surface, and contacts a gelatinous surface called thetectorial membrane. Although hair cells have no axons, they aresurrounded by sensory nerve fibers that form the cochlear branch of thevestibulocochlear nerve (cranial nerve VIII).

As discussed, the oval window, also known as the elliptical windowcommunicates with the stapes to relay sound waves that vibrate from thetympanic membrane. Vibrations transferred to the oval window increasespressure inside the fluid-filled cochlea via the perilymph and scalavestibuli/scala tympani, which in turn causes the round window membraneto expand in response. The concerted inward pressing of the ovalwindow/outward expansion of the round window allows for the movement offluid within the cochlea without a change of intra-cochlear pressure.However, as vibrations travel through the perilymph in the scalavestibuli, they create corresponding oscillations in the vestibularmembrane. These corresponding oscillations travel through the endolymphof the cochlear duct, and transfer to the basilar membrane. When thebasilar membrane oscillates, or moves up and down, the organ of Cortimoves along with it. The hair cell receptors in the Organ of Corti thenmove against the tectorial membrane, causing a mechanical deformation inthe tectorial membrane. This mechanical deformation initiates the nerveimpulse which travels via the vestibulocochlear nerve to the centralnervous system, mechanically transmitting the sound wave received intosignals that are subsequently processed by the central nervous system.

Diseases

Otic disorders, including auris interna disorders, produce symptomswhich include but are not limited to hearing loss, nystagmus, feeling offullness in ear, vertigo, tinnitus, inflammation, and congestion.

Meniere's Disease

Meniere's Disease is an idiopathic condition characterized by suddenattacks of vertigo, nausea and vomiting that may last for 3 to 24 hours,and may subside gradually. Progressive hearing loss, tinnitus and asensation of pressure in the ears accompanies the disease through time.Meniere's disease appears linked to defective fluid homeostasis in theinner ear, including an increase in production or a decrease inreabsorption of inner ear fluid.

The vasopressin (VP) aquaporin 2 (AQP2) system in the inner ear, and inparticular VP, has a role in regulating endolymph production, therebyincreasing pressure in the vestibular and cochlear structures. VP levelsare upregulated in patients with endolymphatic hydrops (Meniere'sDisease), and chronic administration of VP in guinea pigs inducesendolymphatic hydrops. Treatment with VP antagonists, including infusionof OPC-31260 (a competitive antagonist of V₂-R) into the scala tympaniresulted in a marked reduction of Meniere's disease symptoms. Other VPantagonists include WAY-140288, CL-385004, tolvaptan, conivaptan, SR121463A and VPA 985. (Sanghi et al. Eur. Heart J. (2005) 26:538-543;Palm et al. Nephrol. Dial Transplant (1999) 14:2559-2562).

Other treatments are aimed at dealing with the immediate symptoms andprevention of recurrence. Low-sodium diets, avoidance of caffeine,alcohol, and tobacco have been advocated. Medications that temporarilyrelieve vertigo attacks include antihistamines (including meclizine(Antivert®, Bonine®, Dramamine®, Driminate), betahistine and otherantihistamines), and central nervous system agents, includingbarbiturates and/or benzodiazepines, including lorazepam or diazepam.Other examples of drugs that are useful in relieving symptoms includenerve blocking agents (e.g., atropine), muscarinic antagonists,including scopolamine. Nausea and vomiting are relieved by suppositoriescontaining antipsychotic agents, including the phenothiazine agentprochlorperazine (Compazine®, Buccastem, Stemetil and Phenotil).

Surgical procedures that have been used to relieve symptoms include thedestruction of vestibular and/or cochlear function to relieve vertigosymptoms. These procedures aim to either reduce fluid pressure in theinner ear and/or to destroy inner ear balance function. An endolymphaticshunt procedure, which relieves fluid pressure, is placed in the innerear to relieve symptoms of vestibular dysfunction. Other treatmentsinclude gentamicin application, which when injected into the eardrumrelieves fullness in the ear. However, gentamicin destroys sensory haircell function, thereby eradicating inner ear balance function andcausing permanent balance impairment. Severing of the vestibular nervemay also be employed, which while preserving hearing, may controlvertigo.

Meniere's Syndrome

Meniere's Syndrome, which displays similar symptoms as Meniere'sdisease, is attributed as a secondary affliction to an unrelated diseaseprocess, e.g. thyroid disease or inner ear inflammation due to syphilisinfection. Meniere's syndrome, thus, are secondary effects to variousprocess that interfere with normal production or resorption ofendolymph, including endocrine abnormalities, electrolyte imbalance,autoimmune dysfunction, medications, infections (e.g. parasiticinfections) or hyperlipidemia. Treatment of patients afflicted withMeniere's Syndrome is similar to Meniere's Disease.

Vestibular Neuronitis

Vestibular neuronitis is characterized by sudden vertigo attacks, andmay be caused by inflammation of the nerve to the semicircular canalslikely caused by a virus. The first attack of vertigo is usually severe,leading to nystagmus, a condition characterized by the flickering of theeyes involuntarily toward the affected side. Hearing loss does notusually occur. Diagnosis of vestibular neuronitis usually involves testsfor nystagmus using electronystamography, a method of electronicallyrecording eye movements. Magnetic resonance imaging may also beperformed to determine if other causes may play a role in the vertigosymptoms.

Treatment of vertigo is identical to Meniere's disease, and may includemeclizine, lorazepam, prochlorperazine or scopolamine. Fluids andelectrolytes may also be intravenously administered if the vomiting issevere.

Postural Vertigo

Postural vertigo, otherwise known as positional vertigo, ischaracterized by sudden violent vertigo that is triggered by certainhead positions. This condition may be caused by damaged semicircularcanals caused by physical injury to the inner ear, otitis media, earsurgery or blockage of the artery to the inner ear.

Vertigo onset in patients with postural vertigo usually develops when aperson lies on one ear or tilts the head back to look up. Vertigo may beaccompanied by nystagmus. Treatment of postural vertigo involves thesame treatment as in Meniere's disease. In severe cases of posturalvertigo, the vestibular nerve is severed to the affected semicircularcanal.

Ramsay Hunt's Syndrome (Herpes Zoster Infection)

Ramsay Hunt's syndrome is caused by a herpes zoster infection of theauditory nerve. The infection may cause severe ear pain, hearing loss,vertigo, as well as blisters on the outer ear, in the ear canal, and onthe skin of the face or neck supplied by the nerves. Facial muscles mayalso become paralyzed if the facial nerves are compressed by theswelling. Hearing loss may be temporary or permanent, with vertigosymptoms usually lasting from several days to weeks.

Treatment of Ramsay Hunt's syndrome includes administration of antiviralagents, including acyclovir. Other antiviral agents include famciclovirand valacyclovir. Combination of antiviral and corticosteroid therapymay also be employed to ameliorate herpes zoster infection. Analgesicsor narcotics may also be administered to relieve the pain, and diazempamor other central nervous system agents to suppress vertigo. Capsaicin,lidocaine patches and nerve blocks may also be used. Surgery may also beperformed on compressed facial nerves to relieve facial paralysis.

Sensorineural Hearing Loss

Sensorineural hearing loss occurs when the components of the inner earor accompanying neural components are affected, and may contain aneural, i.e. when the auditory nerve or auditory nerve pathways in thebrain are affected, or sensory component. Sensory hearing loss may behereditary, or it may be caused by acoustic trauma (i.e. very loudnoises), a viral infection, drug-induced or Meniere's disease. Neuralhearing loss may occur as a result of brain tumors, infections, orvarious brain and nerve disorders, such as stroke. In some instanceshearing loss is age related hearing loss. Some hereditary diseases, suchas Refsum's disease (defective accumulation of branched fatty acids),may also cause neural disorders affecting hearing loss. Auditory nervepathways may be damaged by demyelinating diseases, e.g. idiopathicinflammatory demyelinating disease (including multiple sclerosis),transverse myelitis, Devic's disease, progressive multifocalleukoencephalopathy, Guillain-Barre syndrome, chronic inflammatorydemyelinating polyneuropathy and anti-MAG perpheral neuropathy.

The incidence of sudden deafness, or sensorineural hearing loss, occursin about 1 in 5000 individuals, and may be caused by viral or bacterialinfections, e.g. mumps, measles, influenza, chickenpox, cytomegalovirus,syphilis or infectious mononucleosis, or physical injury to the innerear organ. In some cases, no cause can be identified. Tinnitus andvertigo may accompany sudden deafness, which subsides gradually. Oralcorticosteroids are frequently prescribed to treat sensorineural hearingloss. In some cases, surgical intervention may be necessary.

Hearing Loss From Excessive Noise

Noise induced hearing loss may occur from prolonged exposure to loudnoises, acoustic trauma such as loud music, heavy equipment ormachinery, airplanes, gunfire or other human-based noises. The hearingloss occurs as result of destruction of hair cell receptors in the innerear. This hearing loss is often accompanied by tinnitus. Permanentdamage to hearing loss is often diagnosed.

Although there is currently no treatment for noise-induced hearing loss,several treatment regimens have been experimentally developed, includingtreatment with insulin-like growth factor 1 (IGF-1). Lee et al. Otol.Neurotol. (2007) 28:976-981).

Hereditary Disorders

Hereditary disorders, including Scheibe, Mondini-Michelle,Waardenburg's, Michel, Alexander's ear deformity, hypertelorism,Jervell-Lange Nielson, Refsum's and Usher's sydromes, are found inapproximately 20% of patients with sensorineural hearing loss.Congenital ear malformations may result from defects in the developmentof the membranous labyrinthine, the osseous labyrinthine, or both. Alongwith profound hearing loss and vestibular function abnormalities,hereditary deformities may also be associated with other dysfunctions,including development of recurring meningitis, cerebral spinal fluid(CSF) leaks, as well as perilymphatic fistulas. Treatment of chronicinfections may be necessitated in hereditary disorder patients.

Autoimmune Inner Ear Disease

Autoimmune inner ear disease (AIED) is one of the few reversible causesof sensorineural hearing loss. It is a rare disorder appearing in bothadults and children that often involves a bilateral disturbance of theaudio and vestibular functions of the auris interna. In many cases, AIEDoccurs without systemic autoimmune symptoms, but up to one-third ofpatients also suffer from a systemic autoimmune illness, such asinflammatory bowel disease, rheumatoid arthritis, Ankylosingspondylitis, Systemic Lupus Erythematosus (SLE), Sjögren's Syndrome,Cogan's disease, ulcerative colitis, Wegener's granulomatosis andscleroderma. Behçet's disease, a multisystem disease, also commonly hasaudiovestibular problems. There is some evidence for food-relatedallergies as a cause for cochlear and vestibular autoimmunity, but thereis presently no agreement as to its importance in the aetiology of thedisease. A classification scheme for AIED has been developed (Harris andKeithley, (2002) Autoimmune inner ear disease, in OtorhinolaryngologyHead and Neck Surgery. 91, 18-32).

The immune system normally performs a crucial role in protecting theinner ear from invasive pathogens such as bacteria and viruses. However,in AIED the immune system itself begins to damage the delicate inner eartissues. The inner ear is fully capable of mounting a localized immuneresponse to foreign antigens. When a foreign antigen enters the innerear, it is first processed by immunocompetent cells which reside in andaround the endolymphatic sac. Once the foreign antigen has beenprocessed by these immunocompetent cells, these cells secrete variouscytokines which modulate the immune response of the inner ear. Oneresult of this cytokine release is to facilitate the influx ofinflammatory cells which are recruited from the systemic circulation.These systemic inflammatory cells enter the cochlea via diapedesisthrough the spiral modiolar vein and its tributaries, and begin toparticipate in antigen uptake and deregulation just as it occurs inother parts of the body. Interleukin 1 (IL-1) plays an important role inmodulating the innate (nonspecific) immune response and is a knownactivator of resting T helper cells and B-cells. T helper cells, onceactivated by IL-1, produce IL-2. IL-2 secretion results indifferentiation of pluripotent T-cells into helper, cytotoxic andsuppressor T-cell subtypes. IL-2 also assists T helper cells in theactivation of B lymphocytes and probably plays a pivotal role in theimmunoregulation of the immune response of the vestibular and cochlearregions. IL-2 is within the perilymph of the auris interna as early as 6h after antigen challenge with peak levels at 18 h after antigenchallenge. The perilymphatic levels of IL-2 then dissipate, and it is nolonger present within the perilymph at 120 hours post antigen challenge.

Pharmaceutical Agents

Provided herein are aural pressure modulating compositions orformulations that ameliorate or lessen otic disorders, includingdisorders of inner ear fluid homeostasis, and their attendant symptoms,which include but are not limited to hearing loss, nystagmus, vertigo,tinnitus, and inflammation. Otic disorders have causes and symptoms thatare responsive to the pharmaceutical agents disclosed herein, or otherpharmaceutical agents. Aural pressure modulating agents which are notdisclosed herein but which are useful for the amelioration oreradication of otic disorders are expressly included and intended withinthe scope of the embodiments presented.

Moreover, pharmaceutical agents which have been previously shown to betoxic, harmful or non-effective during systemic or localized applicationin other organ systems, for example through toxic metabolites formedafter hepatic processing, toxicity of the drug in particular organs,tissues or systems, through high levels needed to achieve efficacy,through the inability to be released through systemic pathways orthrough poor pK characteristics, are useful in some embodiments herein.Accordingly, pharmaceutical agents which have limited or no systemicrelease, systemic toxicity, poor pK characteristics or combinationsthereof are contemplated within the scope of the embodiments disclosedherein.

The aural pressure modulating formulations disclosed herein areoptionally targeted directly to otic structures where treatment isneeded; for example, one embodiment contemplated is the directapplication of the aural pressure modulating formulations disclosedherein onto the round window membrane or the crista fenestrae cochlea ofthe auris interna, allowing direct access and treatment of the aurisinterna, or inner ear components. In other embodiments, the auralpressure modulating formulation disclosed herein is applied directly tothe oval window. In yet other embodiments, direct access is obtainedthrough microinjection directly into the auris interna, for example,through cochlear microperfusion. Such embodiments also optionallycomprise a drug delivery device, wherein the drug delivery devicedelivers the aural pressure modulating formulations through use of aneedle and syringe, a pump, a microinjection device, an in situ formingspongy material or any combination thereof.

Some pharmaceutical agents, either alone or in combination, areototoxic. For example, gentamicin is an antibiotic that is useful intreatment of imbalance in aural pressure but is associated withototoxicity. In some instances, the combination of an ototoxic drug(e.g., an ototoxic aural pressure modulator) in combination with anotoprotectant is protective by lessening the ototoxic effects of thedrug. The localized application of a potentially ototoxic drug alsolessens the toxic effects that otherwise occur through systemicapplication through the use of lower amounts with maintained efficacy,or the use of targeted amounts for a shorter period of time.

Moreover, some pharmaceutical excipients, diluents or carriers arepotentially ototoxic. For example, benzalkonium chloride, a commonpreservative, is ototoxic at certain concentrations and thereforepotentially harmful if introduced into the vestibular or cochlearstructures. In formulating a controlled release aural pressuremodulating formulation, it is advised to avoid or combine theappropriate excipients, diluents or carriers to lessen or eliminatepotential ototoxic components from the formulation, or to decrease theamount of such excipients, diluents or carriers. Optionally, acontrolled release aural pressure modulating formulation includesotoprotective components, such as antioxidants, alpha lipoic acid,calcium, fosfomycin or iron chelators, to counteract potential ototoxiceffects that may arise from the use of specific therapeutic agents orexcipients, diluents or carriers.

Vasopressin and the Vasopressin Receptor

Vasopressin (VP) is a hormone that plays an important part incirculatory and water homoeostasis. This hormone is synthesised byneurosecretory cells located predominantly in two specific hypothalamicnuclei—the supraoptic nucleus and the paraventricular nucleus. Theseneurons have axons that terminate in the neural lobe of the posteriorpituitary gland (neurohypophysis) in which they release vasopressin. Thethree vasopressin receptor subtypes (VP1a, VP1b and VP2) all belong tothe G-protein coupled receptor family and have differing tissuedistributions. The VP1a receptor is predominantly located in thevascular smooth muscle, hepatocytes and blood platelets. The VP1breceptors are found in the anterior pituitary. The VP2 receptors arelocalized in the collecting duct of the kidney and regulate thepresentation of aquaporin-2 channels at the apical cell surface. Theeffect of modulation of the VP2 subtype provides readily observedchanges in urine volume and electrolyte levels to determine thepharmacological effects of anti-diuresis.

Vasopressin regulates systemic osmolality by controlling urinary volumeand composition. Vasopressin is secreted in response to increases inplasma tonicity (very sensitive stimulus) or to decreases in plasmavolume (less sensitive stimulus). Vasopressin mainly regulates urinaryvolume by binding to the VP receptor in the collecting duct of thekidney. The VP receptor also exists in the inner ear of rodents, andaquaporin-2 (AQP2), a VP mediated water channel protein, is alsoexpressed (Kitano et al. Neuroreport (1997), 8:2289-92). Waterhomeostasis of the inner ear fluid was confirmed to be regulated usingthe VP-AQP2 system (Takeda et al. Hear Res (2000), 140:1-6; Takeda etal. Hear Res. (2003), 182:9-18). A recent study looked at tissueexpression of VP2 and AQP2 in human endolymphatic sac byimmunohistochemistry and noted that VP2 and AQP2 were located in theepithelial layer of the endolymphatic sac but not in surroundingconnective tissue (Taguchi et al, Laryngoscope (2007), 117:695-698).Studies on the systemic administration of vasopressio in the guinea pigshowed the development of endolymphatic hydrops (Takeda et al. Hear Res(2000), 140:1-6). Additionally, the aquaporin-4 knockout mouse, whileotherwise healthy, is deaf (Beitz et al., Cellular and MolecularNeurobiology (2003) 23 (3):315-29). This suggests that transport ofwater and solutes in a manner similar to that of the kidney may play arole in fluid homeostasis of the endolymphatic sac.

Vasopressin Receptor Modulators

Vasopressin receptor modulators can be differentiated based upon theirefficacy relative to the vasopressin peptide hormone. A vasopressinreceptor full agonist is a mimic of the native peptide. A vasopressinreceptor antagonist blocks the effect of the native peptide. A partialagonist can serve as a mimic of the native peptide and induce a partialresponse, or in the presence of elevated levels of the native peptide, apartial agonist competes with the native peptide for receptor occupancyand provides a reduction in efficacy, relative to the native peptidealone. For a vasopressin receptor with constitutive activity, an inverseagonist serves to reverse the activity of the receptor.

Agonists of the VP2 receptor are known, including OPC-51803 and relatedanalogs (Kondo et al., J. Med. Chem. (2000) 43:4388; Nakamura et al.,Br. J. Pharmacol. (2000) 129 (8):1700; Nakamure et al., J. Pharmacol.Exp. Ther. (2000) 295 (3):1005) and WAY-VNA-932 (Caggiano, Drugs Gut(2002) 27 (3):248). Antagonists of the VP2 receptor include lixivaptan,tolvaptan, conivaptan, SR-121463 and OPC-31260 (Martin et al., J. Am.Soc. Nephrol. (1999) 10 (10):2165; Gross et al., Exp. Physiol. (2000)85: Spec No 2.53S; Wong et al., Gastroent April 2000, vol 118, 4 Suppl.2, Part 1); Norman et al., Drugs Fut. (2000), 25 (11):1121; Inoue etal., Clin. Pharm. Therap. (1998) 63 (5); 561). In testing against theconstitutively activated D136A mutant VP2 receptor, SR-1211463 andOPC-31260 behaved as inverse agonist (Morin et al., FEBS Letters (1998)441 (3):470-75).

Prostaglandins

Prostaglandins are members of a group of fatty-acid derived compoundsand depending upon the subtype, participate in a variety of functions,including control of constriction or dilation in vascular smooth musclecells, aggregation or disaggregation of platelets, sensitization ofspinal neurons to pain, increase or decrease in intraocular pressure,regulation of inflammatory mediation, regulation of calcium movement,control of hormone regulation and control of hormonal regulation.Prostaglandins have both paracrine and autocrine functions, and are asubclass of eicosanoid compounds.

Prostaglandin analogues, such as latanoprost, travoprost, unoprostone,minprostin F2 alpha and bimtoprost, have been shown to reduceintra-ocular pressure in glaucoma patients by enhancing the uveoscleraloutflow, possibly through vasodilation mechanisms, in addition toeffects on the trabecular meshwork. In sensorineural hearing loss animalmodels, noise exposure induces 8-isoprostaglandin F2α production in thecochlea, concomitant with an increase in vasoconstriction and reducedblood flow. Treatment with SQ29548, a specific antagonist of8-isoprostaglandin F2α, prevents these noise-induced changes in cochlearblood flow and vascular conductance. Inhibition of prostaglandin F2αfunction also reduces tinnitus in patients suffering from Meniere'sdisease, as well as improvements in hearing and vertigo. Finally,prostaglandins have been implicated in chronic inflammation associatedwith otitis media.

Accordingly, one embodiment disclosed herein is the use of prostaglandinmodulators, including latanoprost, travoprost, unoprostone, minprostinF2-alpha, bimtoprost and SQ29548, to ameliorate or decrease inner earand middle ear disorders, including Meniere's disease, tinnitus,vertigo, hearing loss and otitis media.

Estrogen-Related Receptor Beta Modulators

Estrogen-related receptor beta (ERR-beta; also known as Nr3b2), anorphan nuclear receptor, is specifically expressed in and controls thedevelopment of the endolymph-producing cells of the inter ear: thestrial marginal cells in the cochlea and the vestibular dark cells inthe ampulla and utricle. (Chen et al. Dev. Cell. (2007) 13:325-337).Nr3b2 expression has been localized in the endolymph-secreting strialmarginal cells and vestibular dark cells of the cochlea and vestibularapparatus, respectively. Studies in knockout mice have shown that strialmarginal cells in these animals fail to express multiple ion channel andtransporter genes, suggesting a role in the development and/or functionof endolymph producing epithelia. Moreover, conditional knockout of theNr3b2 gene results in deafness and diminished endolymphatic fluidvolume.

Other studies suggest a role for estrogen-related receptor β/NR3B2(ERR/Nr3b2) in regulating endolymph production, and therefore pressurein the vestibular/cochlear apparatus. Treatment with antagonists toERR/Nr3b2 may assist in reducing endolymphatic volume, and thus alterpressure in the auris interna structures. Accordingly, agents whichantagonize ERR/Nr3b2 expression, protein production or protein functionare contemplated as useful with the formulations disclosed herein.

Osmotic Diuretics

Contemplated for use with the compositions disclosed herein, are agentswhich regulate aural pressure. Accordingly, some embodiments compriseosmotic diuretics. An osmotic diuretic is a substance that produces anosmotic gradient between two spaces. In certain instances, an osmoticdiuretic produces an osmotic gradient between the endolymphatic andperilymphatic spaces. In certain instances, an osmotic gradient betweenthe endolymphatic and perilymphatic spaces exerts a dehydrating effecton the endolymphatic space. In certain instances, dehydrating theendolymphatic space decreases aural pressure.

Accordingly, in some embodiments of the compositions and formulationsdisclosed herein, the aural pressure modulator is an osmotic diuretic.In some embodiments, the osmotic diuretic is erythritol, mannitol,glucose, isosorbide, glycerol; urea; or combinations thereof.

Calcium Channel Blockers

Contemplated for use with the formulations disclosed herein are agentsthat modulate fluid homeostasis in the ear. Accordingly, someembodiments incorporate the use of ion channel modulators (e.g.,potassium, sodium or calcium channel modulators, Na/K ATPase modulators,or KCNQ channel modulators) in the compositions described herein. Insome instances, Ca²⁺ concentration is critical for cochlear homeostasisand sensory transduction in the cochlea. Calcium channels are channelsformed in the plasma membrane of neurons (amongst other cells) byintegral membrane proteins. These channels conduct Ca⁺ through a cell'splasma membrane. In neurons, the flow of Ca²⁺ is partly responsible forcreating and propagating action potentials in neurons. It can also beresponsible for the release of certain neurotransmitters.

In some embodiments, calcium channel antagonists modulate fluidhomeostasis in the ear. In some embodiments, the calcium channelantagonist is lomerizine, cinnarizine, flunarizine, or nimodipine. Insome embodiments, the calcium channel antagonist is cinnarizine. In someembodiments, the calcium channel antagonist is flunarizine. In someembodiments, the calcium channel antagonist is nimodipine. In someembodiments, the calcium channel antagonist is lomerizine.

Antibiotics

Contemplated for use with the formulations disclosed herein are agentsthat alleviate a feeling of fullness in the ear. Accordingly, someembodiments incorporate the use of antibacterial agents including, butnot limited to, amikacin, gentamicin, kanamycin, neomycin, netilmicin,streptomycin, tobramycin, paromomycin, geldanmycin, herbimycin,loracarbef, ertapenem, doripenem, imipenem, cilastatin, meropenem,cefadroxil, cefazolin, cefalotin, cefalexin, cefaclor, cefamandole,cefoxitin, defprozil, cefuroxime, cefixime, cefdinir, cefditoren,cefoperazone, cefotaxime, cefpodoxime, ceftazidime, ceftibuten,ceftizoxime, ceftriaxone, cefepime, ceftobiprole, teicoplanin,vancomycin, azithromycin, clarithromycin, dirithromycin, erythromycin,roxithromycin, troleandomycin, telithromycin, spectinomycin, aztreonam,amoxicillin, ampicillin, azlocillin, carbenicillin, cloxacillin,dicloxacillin, flucloxacillin, mezlocillin, meticillin, nafcillin,oxacillin, penicillin, piperacillin, ticarcillan, bacitracin, colistin,polymyxin B, ciprofloxacin, enoxacin, gatifloxacin, levofloxacin,lomefloxacin, moxifloxacin, norfloxacin, ofloxacin, trovfloxacin,mafenide, prontosil, sulfacetamide, sulfamethizole, sulfanimilimde,sulfsalazine, sulfsioxazole, trimethoprim, demeclocycline, doxycycline,minocycline, oxtetracycline, tetracycline, arsphenamine,chloramphenicol, clindamycin, lincomycin, ethambutol, fosfomycin,fusidic acid, furazolidone, isoniazid, linezolid, metronidazole,mupirocin, nitrofurantoin, platensimycin, pyrazinamide,quinuspristin/dalfopristin, rifampin, tinidazole, and combinationsthereof.

RNAi

In some embodiments, where inhibition or down-regulation of a target isdesired (e.g. genes ERR, and Nr3b2), RNA interference may be utilized.In some embodiments, the agent that inhibits or down-regulates thetarget is an siRNA molecule. In certain instances, the siRNA moleculeinhibits the transcription of a target by RNA interference (RNAi). Insome embodiments, a double stranded RNA (dsRNA) molecule with sequencescomplementary to a target is generated (e.g by PCR). In someembodiments, a 20-25 bp siRNA molecule with sequences complementary to atarget is generated. In some embodiments, the 20-25 bp siRNA moleculehas 2-5 bp overhangs on the 3′ end of each strand, and a 5′ phosphateterminus and a 3′ hydroxyl terminus. In some embodiments, the 20-25 bpsiRNA molecule has blunt ends. For techniques for generating RNAsequences see Molecular Cloning: A Laboratory Manual, second edition(Sambrook et al., 1989) and Molecular Cloning: A Laboratory Manual,third edition (Sambrook and Russel, 2001), jointly referred to herein as“Sambrook”); Current Protocols in Molecular Biology (F. M. Ausubel etal., eds., 1987, including supplements through 2001); Current Protocolsin Nucleic Acid Chemistry John Wiley & Sons, Inc., New York, 2000) whichare hereby incorporated by reference for such disclosure.

In some embodiments, the dsRNA or siRNA molecule is incorporated into acontrolled-release auris-acceptable microsphere or microparticle,hydrogel, liposome, or thermoreversible gel. In some embodiments, theauris-acceptable microsphere, hydrogel, liposome, paint, foam, in situforming spongy material, nanocapsule or nanosphere or thermoreversiblegel is injected into the inner ear. In some embodiments, theauris-acceptable microsphere or microparticle, hydrogel, liposome, orthermoreversible gel. In some embodiments, the auris-acceptablemicrosphere, hydrogel, liposome, paint, foam, in situ forming spongymaterial, nanocapsule or nanosphere or thermoreversible gel is injectedinto the cochlea, the organ of Corti, the vestibular labyrinth, or acombination thereof.

In certain instances, after administration of the dsRNA or siRNAmolecule, cells at the site of administration (e.g. the cells ofcochlea, organ of Corti, and/or the vestibular labyrinth) aretransformed with the dsRNA or siRNA molecule. In certain instancesfollowing transformation, the dsRNA molecule is cleaved into multiplefragments of about 20-25 bp to yield siRNA molecules. In certaininstances, the fragments have about 2 bp overhangs on the 3′ end of eachstrand.

In certain instances, an siRNA molecule is divided into two strands (theguide strand and the anti-guide strand) by an RNA-induced SilencingComplex (RISC). In certain instances, the guide strand is incorporatedinto the catalytic component of the RISC (i.e. argonaute). In certaininstances, the guide strand binds to a complementary target mRNAsequence. In certain instances, the RISC cleaves the target mRNA. Incertain instances, the expression of the target gene is down-regulated.

In some embodiments, a sequence complementary to a target is ligatedinto a vector. In some embodiments, the sequence is placed between twopromoters. In some embodiments, the promoters are orientated in oppositedirections. In some embodiments, the vector is contacted with a cell. Incertain instances, a cell is transformed with the vector. In certaininstances following transformation, sense and anti-sense strands of thesequence are generated. In certain instances, the sense and anti-sensestrands hybridize to form a dsRNA molecule which is cleaved into siRNAmolecules. In certain instances, the strands hybridize to form an siRNAmolecule. In some embodiments, the vector is a plasmid (e.g pSUPER;pSUPER.neo; pSUPER.neo+gfp).

In some embodiments, the vector is incorporated into acontrolled-release auris-acceptable microsphere or microparticle,hydrogel, liposome, or thermoreversible gel. In some embodiments, theauris-acceptable microsphere, hydrogel, xerogel, liposome, paint, foam,in situ forming spongy material, nanocapsule or nanosphere orthermoreversible gel is injected into the inner ear. In someembodiments, the auris-acceptable microsphere or microparticle,hydrogel, liposome, actinic radiation curable gel, or thermoreversiblegel. In some embodiments, the auris-acceptable microsphere, hydrogel,liposome, paint, foam, in situ forming spongy material, nanocapsule ornanosphere or thermoreversible gel is injected into the cochlea, theorgan of Corti, the vestibular labyrinth, or a combination thereof.

In some embodiments, the compositions described herein have aconcentration of active pharmaceutical ingredient between about 0.01% toabout 90%, between about 0.01% to about 50%, between about 0.1% to about70%, between about 0.1% to about 50%, between about 0.1% to about 40%,between about 0.1% to about 30%, between about 0.1% to about 20%,between about 0.1% to about 10%, or between about 0.1% to about 5%, ofthe active ingredient, or pharmaceutically acceptable prodrug or saltthereof, by weight of the composition. In some embodiments, thecompositions described herein have a concentration of activepharmaceutical agent between about 1% to about 50%, between about 5% toabout 50%, between about 10% to about 40%, or between about 10% to about30%, of the active ingredient, or pharmaceutically acceptable prodrug orsalt thereof, by weight of the composition. In some embodiments,formulations described herein comprise about 70% by weight of an auralpressure modulating agent by weight of the formulation. In someembodiments, formulations described herein comprise about 60% by weightof an aural pressure modulating agent by weight of the formulation. Insome embodiments, formulations described herein comprise about 50% byweight of an aural pressure modulating agent by weight of theformulation. In some embodiments, formulations described herein compriseabout 40% by weight of an aural pressure modulating agent by weight ofthe formulation. In some embodiments, formulations described hereincomprise about 30% by weight of an aural pressure modulating agent byweight of the formulation. In some embodiments, formulations describedherein comprise about 20% by weight of an aural pressure modulatingagent by weight of the formulation. In some embodiments, formulationsdescribed herein comprise about 15% by weight of an aural pressuremodulating agent, or pharmaceutically acceptable prodrug or saltthereof, by weight of the formulation. In some embodiments, formulationsdescribed herein comprise about 10% by weight of an aural pressuremodulating agent by weight of the formulation. In some embodiments,formulations described herein comprise about 5% by weight of an auralpressure modulating agent, or pharmaceutically acceptable prodrug orsalt thereof, by weight of the formulation. In some embodiments,formulations described herein comprise about 2.5% by weight of an auralpressure modulating agent, or pharmaceutically acceptable prodrug orsalt thereof, by weight of the formulation. In some embodiments,formulations described herein comprise about 1% by weight of an auralpressure modulating agent, or pharmaceutically acceptable prodrug orsalt thereof, by weight of the formulation. In some embodiments,formulations described herein comprise about 0.5% by weight of an auralpressure modulating agent, or pharmaceutically acceptable prodrug orsalt thereof, by weight of the formulation. In some embodiments,formulations described herein comprise about 0.1% by weight of an auralpressure modulating agent, or pharmaceutically acceptable prodrug orsalt thereof, by weight of the formulation. In some embodiments,formulations described herein comprise about 0.01% by weight of an auralpressure modulating agent, or pharmaceutically acceptable prodrug orsalt thereof, by weight of the formulation. In some embodiments, theformulations described herein have a concentration of activepharmaceutical ingredient, or pharmaceutically acceptable prodrug orsalt thereof, between about 0.1 to about 70 mg/mL, between about 0.5mg/mL to about 70 mg/mL, between about 0.5 mg/mL to about 50 mg/mL,between about 0.5 mg/mL to about 20 mg/mL, between about 1 mg to about70 mg/mL, between about 1 mg to about 50 mg/mL, between about 1 mg/mLand about 20 mg/mL, between about 1 mg/mL to about 10 mg/mL, or betweenabout 1 mg/mL to about 5 mg/mL, of the active agent, or pharmaceuticallyacceptable prodrug or salt thereof, by volume of the formulation.

Combination Therapy

In some embodiments, the compositions disclosed herein further comprisean additional therapeutic agent. In some embodiments, the additionaltherapeutic agent is a diuretic, an anti-TNF agent, an anti-emeticagent, an anti-vertigo agent, a local acting anesthetic agent, ananti-anxiety agent, a corticosteroid, an anti-histamine, or combinationsthereof. In some embodiments, the additional therapeutic agent is anotoprotectant, a Na/K ATPase modulator, a KCNQ channel modulator, orcombinations thereof. Any combination of one or more pharmaceuticalagents (e.g., aural pressure modulators) and one or more additionaltherapeutic agents is compatible with the compositions and devicesdescribed herein.

Diuretic Agents

Contemplated for use in combination with the aural pressure modulatingformulations disclosed herein are diuretic agents. A diuretic agent is adrug that elevates the rate of urination. Such diuretics includetriamterene, amiloride, bendroflumethiazide, hydrochlorothiazide,furosemide, torsemide, bumetanide, acetazolamide, dorzolamide andcombinations thereof.

Anti-TNF Agents

Contemplated for use in combination with the aural pressure modulatingformulations disclosed herein are anti-TNF agents which reduce orameliorate symptoms or effects (e.g., aural pressure) as a result of anautoimmune disease and/or inflammatory disorder, including AIED.Autoimmune deficiencies may be a contributing factor to otic disorderssuch as Meniere's disease. Accordingly, some embodiments incorporate theuse of agents which block the effects of TNF-α, including anti-TNFagents. By way of example only, anti-TNF agents include etanercept(ENBREL®), infliximab (REMICADE®), adalimumab (HUMIRA®), and golimumab(CNTO 148) or combinations thereof. Infliximab and adalimumab areanti-TNF monoclonal antibodies, and etanercept is a fusion proteindesigned to bind specifically to the TNF protein. All are currentlyapproved for use in the treatment of rheumatoid arthritis. Golimumab,which is currently in Phase 3 clinical trials for rheumatoid arthritis,psoriatic arthritis and ankylosing spondylitis, is a fully-humanizedanti-TNF-alpha IgG1 monoclonal antibody that targets and neutralizesboth the soluble and the membrane-bound form of TNF. Other antagoniststo TNF, by way of example only, include TNF receptors (pegylated solubleTNF receptor type 1; Amgen); TNF binding factors (Onercept; Serono); TNFantibodies (US Patent App. No. 2005/0123541; US Patent App. No.2004/0185047); single domain antibodies against the p55 TNF receptor (USPatent App. No. 2008/00088713); soluble TNF receptors (US Patent App.No. 2007/0249538); fusion polypeptides binding to TNF (US Patent App.No. 2007/0128177); TNF-α converting enzyme inhibitors (Skotnicki et al,Annual Reports in Medicinal Chemistry (2003), 38, 153-162); IKKinhibitors (Karin et al, Nature Reviews Drug Discovery (2004), 3, 17-26)and flavone derivatives (US Patent App. No. 2006/0105967), all of whichare incorporated by reference for such disclosure. The use of onercept,a soluble TNF p55 receptor, was discontinued in 2005. Three phase-IIIclinical trials reported patients diagnosed with fatal sepsis. A risk tobenefit analysis was subsequently performed, resulting in thediscontinuation of the clinical trials. As discussed above, theembodiments herein specifically contemplate the use of anti-TNF agentswhich have been previously shown to have limited or no systemic release,systemic toxicity, poor pK characteristics of combinations thereof.

It is contemplated that the localized application of the anti-TNF agentsin combination with aural pressure modulators to the target oticstructures for treatment of autoimmune and/or inflammatory disorderswill result in the reduction or elimination of these adverse sideeffects experienced with systemic treatment. Moreover, localizedtreatment with the anti-TNF agents contemplated herein will also reducethe amount of agent needed for effective treatment of the targeteddisorder due, for example, to the existence of the biological bloodbarrier in the auris interna.

Corticosteroids

Contemplated for use in combination with the aural pressure modulatingagent formulations disclosed herein are corticosteroid agents whichreduce or ameliorate symptoms or effects (e.g., sensation of fullness inthe ear) as a result of an autoimmune disease and/or inflammatorydisorder, including AIED. Such autoimmune response may be a contributingfactor to otic disorders such as Meniere's disease. In some instances,corticosteroids alleviate aural fullness. In some embodiments,corticosteroids are contemplated as monotherapy. Such steroids includeprednisolone, dexamethasone, dexamethasone phosphate, beclomethasone,21-acetoxypregnenolone, alclometasone, algestone, amcinonide,beclomethasone, betamethasone, budesonide, chloroprednisone, clobetasol,clobetasone, clocortolone, cloprednol, corticosterone, cortisone,cortivazol, deflazacort, desonide, desoximetasone, diflorasone,diflucortolone, difluprednate, enoxolone, fluazacort, flucloronide,flumethasone, flunisolide, fluocinolone acetonide, fluocinonide,fluocortin butyl, fluocortolone, fluorometholone, fluperolone acetate,fluprednidene acetate, fluprednisolone, flurandrenolide, fluticasonepropionate, formocortal, halcinonide, halobetasol propionate,halometasone, halopredone acetate, hydrocortamate, hydrocortisone,loteprednol etabonate, mazipredone, medrysone, meprednisone,methylprednisolone, mometasone furoate, paramethasone, prednicarbate,prednisolone, prednisolone 25-diethylamino-acetate, prednisolone sodiumphosphate, prednisone, prednival, prednylidene, rimexolone, tixocortol,triamcinolone, triamcinolone acetonide, triamcinolone benetonide,triamcinolone hexacetonide and combinations thereof.

Anti-Emetic Agents/Central Nervous System Agents

Anti-Emetic agents are optionally used in combination with the auralpressure modulator formulations disclosed herein. Anti-emetic agentsinclude antihistamines and central nervous agents, includinganti-psychotic agents, barbiturates, benzodiazepines and phenothiazines.Other anti-emetic agents include the serotonin receptor antagonists,which include dolasetron, granisetron, ondansetron, tropisetron,palonosetron, and combinations thereof; dopamine antagonists, includingdomperidone, properidol, haloperidol, chlorpromazine, promethazine,prochlorperazine and combinations thereof; cannabinoids, includingdronabinol, nabilone, sativex, and combinations thereof;anticholinergics, including scopolamine; and steroids, includingdexamethasone; trimethobenzamine, emetrol, propofol, muscimol, andcombinations thereof.

Optionally, Central Nervous System agents and barbiturates are useful inthe treatment of nausea and vomiting symptoms that accompany an oticdisorder. When used, an appropriate barbiturate and/or central nervoussystem agent is selected to relieve or ameliorate specific symptomswithout possible side effects, including ototoxicity. Moreover, asdiscussed above, targeting of the drugs to the round window membrane ofthe auris interna reduces possible side effects and toxicity caused bysystemic administration of these drugs. Barbiturates, which act as acentral nervous system depressant, include allobarbital, alphenal,amobarbital, aprobarbital, barnexaclone, barbital, brallobarbital,butabarbital, butalbital, butallylonal, butobarbital, corvalol,crotylbarbital, cyclobarbital, cyclopal, ethallobarbital, febarbamate,heptabarbital, hexethal, hexobarbital, metharbital, methohexital,methylphenobarbital, narcobarbital, nealbarbital, pentobarbital,phenobarbital, primidone, probarbital, propallylonal, proxibarbital,reposal, secobarbital, sigmodal, sodium thiopental, talbutal,thialbarbital, thiamylal, thiobarbital, thiobutabarbital, tuinal,valofane, vinbarbital, vinylbital, and combinations thereof.

Other central nervous system agents which are optionally used inconjunction with the aural pressure modulator formulations disclosedherein include benzodiazepines or phenothiazines. Useful benzodiazepinesinclude, but are not limited to diazepam, lorazepam, oxazepam, prazepam,alprazolam, bromazepam, chlordiazepoxide, clonazepam, clorazepate,brotizolam, estazolam, flunitrazepam, flurazepam, loprazolam,lormetazepam, midazolam, nimetazepam, nitrazepam, ternazepam, triazolam,and combinations thereof. Examples of phenothiazines includeprochlorperazine, chlorpromazine, promazine, triflupromazine,levopromazine, methotrimepramazine, mesoridazine, thiroridazine,fluphenazine, perphenazine, flupentixol, trifluoperazine, andcombinations thereof. In some embodiments, nerve blocking agents (e.g.,atropine) are used in conjunction with the aural pressure modulatorformulations disclosed herein. In some instances, CNS modulators relievepressure in the ear.

Antihistamines, or histamine antagonists, act to inhibit the release oraction of histamine. Antihistamines that target the H1 receptor areuseful in the alleviation or reduction of nausea and vomiting symptomsthat are associated with otic disorders. Such antihistamines include,but are not limited to, meclizine, diphenhydramine, loratadine andquetiapine. Other antihistamines include mepyramine, piperoxan,antazoline, carbinoxamine, doxylamine, clemastine, dimenhydrinate,pheniramine, chlorphenamine, chlorpheniramine, dexchlorpheniramine,brompheniramine, triprolidine, cyclizine, chlorcyclizine, hydroxyzine,promethazine, alimemazine, trimeprazine, cyproheptadine, azatadine,ketotifen, oxatomide and combinations thereof.

Otoprotectants

In some embodiments, any otic formulation described herein furthercomprises otoprotectants that reduce, inhibit or ameliorate theototoxicity of aural pressure modulating agents such as vasopressinmodulators or antibiotics as described herein. Examples ofotoprotectants include, and are not limited to, thiols and/or thiolderivatives and/or pharmaceutically acceptable salts, or derivatives(e.g. prodrugs) thereof (e.g., D-methionine, L-methionine, ethionine,hydroxyl methionine, methioninol, amifostine, mesna (sodium2-sulfanylethanesulfonate), a mixture of D and L methionine,normethionine, homomethionine, S-adenosyl-L-methionine),diethyldithiocarbamate, ebselen(2-phenyl-1,2-benzisoselenazol-3(2H)-one), sodium thiosulfate, AM-111 (acell permeable JNK inhibitor, (Laboratoires Auris SAS)), leucovorin,leucovorin calcium, dexrazoxane, piracetam, Oxiracetam, Aniracetam,Pramiracetam, Phenylpiracetam (Carphedon), Etiracetam, Levetiracetam,Nefiracetam, Nicoracetam, Rolziracetam, Nebracetam, Fasoracetam,Coluracetam, Dimiracetam, Brivaracetam, Seletracetam, Rolipramand orcombinations thereof. Otoprotectants allow for the administration ofotic agents (e.g., aural pressure modulators described herein) at dosesthat are higher than conventional doses; the otic agents would otherwisebe systemically administered at lower doses because of associatedototoxicity.

Presented below (Table 1) are examples of active agents contemplated foruse with the formulations and devices disclosed herein. One or moreactive agents are used in any of the formulations or devices describedherein.

Active Agents (including pharmaceutically acceptable salts of theseactive agents) for use with the Formulations Disclosed Herein

TABLE 1 Auris Condition Therapeutic Agent Benign ParoxysmalDiphenhydramine Positional Vertigo Benign Paroxysmal LorazepamPositional Vertigo Benign Paroxysmal Meclizine Positional Vertigo BenignParoxysmal Oldansetron Positional Vertigo Hearing Loss Estrogen AIEDlixivaptan AIED tovaptan Hearing Loss Estrogen and progesterone (E + P)Hearing Loss Folic acid Hearing Loss Lactated Ringer's with 0.03%Ofloxacin Hearing Loss Methotrexate Hearing Loss N-acetyl cysteineMeniere's Disease Betahistine Meniere's Disease Sildenafil Meniere'sDisease conivaptan Middle Ear Effusion Pneumonococcal vaccine OtitisExterna Diclofenac sodium; dexotc Otitis Externa, AcuteAL-15469A/AL-38905 Otitis Media Amoxicillin/clavulanate Otitis MediaDornase alfa Otitis Media Echinacea purpurea Otitis Media Faropenemmedoxomil Otitis Media Levofloxacin Otitis Media PNCRM9 Otitis MediaPneumococcal vaccine Otitis Media Telithromycin Otitis Media Zmax OtitisMedia with Lansoprazole Effusion Otitis Media, Acute AL-15469A; AL-38905Otitis Media, Acute Amoxicillin Otitis Media, AcuteAmoxicillin-clavulanate Otitis Media, Acute Azithromycin Otitis Media,Acute Azithromycin SR Otitis Media, Acute Cefdinir Otitis Media, AcuteHyland's earache drops Otitis Media, Acute Montelukast Otitis Media,Acute Pneumonococcal vaccine Otitis Media, Acute AL-15469A/AL38905 withTypanostomy Tubes Otitis Media, Chronic Sulfamethoxazole- trimethoprimOtitis Media, Azithromycin Suppurative Otitis Media, TelithromycinSuppurative Otosclerosis Acetylcysteine Ototoxicity Aspirin TinnitusAcamprosate Tinnitus Gabapentin Tinnitus Modafinil Tinnitus NeramexaneTinnitus Neramexane mesylate Tinnitus Piribedil Tinnitus VardenafilTinnitus Vestipitant + Paroxetine Tinnitus Vestiplitant Tinnitus ZincsulfateGeneral Methods of Sterilization

Provided herein are otic compositions that ameliorate or lessen oticdisorders described herein. Further provided herein are methodscomprising the administration of said otic compositions. In someembodiments, the compositions or devices are sterilized. Included withinthe embodiments disclosed herein are means and processes forsterilization of a pharmaceutical composition or device disclosed hereinfor use in humans. The goal is to provide a safe pharmaceutical product,relatively free of infection causing micro-organisms. The U.S. Food andDrug Administration has provided regulatory guidance in the publication“Guidance for Industry: Sterile Drug Products Produced by AsepticProcessing” available at: http://www.fda.gov/cder/guidance/5882fnl.htm,which is incorporated herein by reference in its entirety.

As used herein, sterilization means a process used to destroy or removemicroorganisms that are present in a product or packaging. Any suitablemethod available for sterilization of objects and compositions is used.Available methods for the inactivation of microorganisms include, butare not limited to, the application of extreme heat, lethal chemicals,or gamma radiation. In some embodiments is a process for the preparationof an otic therapeutic formulation comprising subjecting the formulationto a sterilization method selected from heat sterilization, chemicalsterilization, radiation sterilization or filtration sterilization. Themethod used depends largely upon the nature of the device or compositionto be sterilized. Detailed descriptions of many methods of sterilizationare given in Chapter 40 of Remington: The Science and Practice ofPharmacy published by Lippincott, Williams & Wilkins, and isincorporated by reference with respect to this subject matter.

Sterilization by Heat

Many methods are available for sterilization by the application ofextreme heat. One method is through the use of a saturated steamautoclave. In this method, saturated steam at a temperature of at least121° C. is allowed to contact the object to be sterilized. The transferof heat is either directly to the microorganism, in the case of anobject to be sterilized, or indirectly to the microorganism by heatingthe bulk of an aqueous solution to be sterilized. This method is widelypracticed as it allows flexibility, safety and economy in thesterilization process.

Dry heat sterilization is a method which is used to kill microorganismsand perform depyrogenation at elevated temperatures. This process takesplace in an apparatus suitable for heating HEPA-filteredmicroorganism-free air to temperatures of at least 130-180° C. for thesterilization process and to temperatures of at least 230-250° C. forthe depyrogenation process. Water to reconstitute concentrated orpowdered formulations is also sterilized by autoclave. In someembodiments, the formulations described herein comprise micoronizedaural pressure modulating agents (e.g., micro-lixivaptan) that aresterilized by dry heating, e.g., heating for about 7-11 hours atinternal powder temperatures of 130-140° C., or for 1-2 hours atinternal temperatures of 150-180° C.

Chemical Sterilization

Chemical sterilization methods are an alternative for products that donot withstand the extremes of heat sterilization. In this method, avariety of gases and vapors with germicidal properties, such as ethyleneoxide, chlorine dioxide, formaldehyde or ozone are used as theanti-apoptotic agents. The germicidal activity of ethylene oxide, forexample, arises from its ability to serve as a reactive alkylatingagent. Thus, the sterilization process requires the ethylene oxidevapors to make direct contact with the product to be sterilized.

Radiation Sterilization

One advantage of radiation sterilization is the ability to sterilizemany types of products without heat degradation or other damage. Theradiation commonly employed is beta radiation or alternatively, gammaradiation from a ⁶⁰Co source. The penetrating ability of gamma radiationallows its use in the sterilization of many product types, includingsolutions, compositions and heterogeneous mixtures. The germicidaleffects of irradiation arise from the interaction of gamma radiationwith biological macromolecules. This interaction generates chargedspecies and free radicals. Subsequent chemical reactions, such asrearrangements and cross-linking processes, result in the loss of normalfunction for these biological macromolecules. The formulations describedherein are also optionally sterilized using beta irradiation.

Filtration

Filtration sterilization is a method used to remove but not destroymicroorganisms from solutions. Membrane filters are used to filterheat-sensitive solutions. Such filters are thin, strong, homogenouspolymers of mixed cellulosic esters (MCE), polyvinylidene fluoride (PVF;also known as PVDF), or polytetrafluoroethylene (PTFE) and have poresizes ranging from 0.1 to 0.22 μm. Solutions of various characteristicsare optionally filtered using different filter membranes. For example,PVF and PTFE membranes are well suited to filtering organic solventswhile aqueous solutions are filtered through PVF or MCE membranes.Filter apparatus are available for use on many scales ranging from thesingle point-of-use disposable filter attached to a syringe up tocommercial scale filters for use in manufacturing plants. The membranefilters are sterilized by autoclave or chemical sterilization.Validation of membrane filtration systems is performed followingstandardized protocols (Microbiological Evaluation of Filters forSterilizing Liquids, Vol 4, No. 3. Washington, D.C: Health IndustryManufacturers Association, 1981) and involve challenging the membranefilter with a known quantity (ca. 10⁷/cm²) of unusually smallmicroorganisms, such as Brevundimonas diminuta (ATCC 19146).

Pharmaceutical compositions are optionally sterilized by passing throughmembrane filters. Formulations comprising nanoparticles (U.S. Pat. No.6,139,870) or multilamellar vesicles (Richard et al., InternationalJournal of Pharmaceutics (2006), 312 (1-2):144-50) are amenable tosterilization by filtration through 0.22 μm filters without destroyingtheir organized structure.

In some embodiments, the methods disclosed herein comprise sterilizingthe formulation (or components thereof) by means of filtrationsterilization. In another embodiment the auris-acceptable otictherapeutic agent formulation comprises a particle wherein the particleformulation is suitable for filtration sterilization. In a furtherembodiment said particle formulation comprises particles of less than300 nm in size, of less than 200 nm in size, of less than 100 nm insize. In another embodiment the auris-acceptable formulation comprises aparticle formulation wherein the sterility of the particle is ensured bysterile filtration of the precursor component solutions. In anotherembodiment the auris-acceptable formulation comprises a particleformulation wherein the sterility of the particle formulation is ensuredby low temperature sterile filtration. In a further embodiment, lowtemperature sterile filtration is carried out at a temperature between 0and 30° C., between 0 and 20° C., between 0 and 10° C., between 10 and20° C., or between 20 and 30° C.

In another embodiment is a process for the preparation of anauris-acceptable particle formulation comprising: filtering the aqueoussolution containing the particle formulation at low temperature througha sterilization filter; lyophilizing the sterile solution; andreconstituting the particle formulation with sterile water prior toadministration. In some embodiments, a formulation described herein ismanufactured as a suspension in a single vial formulation containing themicronized active pharmaceutical ingredient. A single vial formulationis prepared by aseptically mixing a sterile poloxamer solution withsterile micronized active ingredient (e.g., lixivaptan) and transferringthe formulation to sterile pharmaceutical containers. In someembodiments, a single vial containing a formulation described herein asa suspension is resuspended before dispensing and/or administration.

In specific embodiments, filtration and/or filling procedures arecarried out at about 5° C. below the gel temperature (Tgel) of aformulation described herein and with viscosity below a theoreticalvalue of 100 cP to allow for filtration in a reasonable time using aperistaltic pump.

In another embodiment the auris-acceptable otic therapeutic agentformulation comprises a nanoparticle formulation wherein thenanoparticle formulation is suitable for filtration sterilization. In afurther embodiment the nanoparticle formulation comprises nanoparticlesof less than 300 nm in size, of less than 200 nm in size, or of lessthan 100 nm in size. In another embodiment the auris-acceptableformulation comprises a microsphere formulation wherein the sterility ofthe microsphere is ensured by sterile filtration of the precursororganic solution and aqueous solutions. In another embodiment theauris-acceptable formulation comprises a thermoreversible gelformulation wherein the sterility of the gel formulation is ensured bylow temperature sterile filtration. In a further embodiment, the lowtemperature sterile filtration occurs at a temperature between 0 and 30°C., or between 0 and 20° C., or between 0 and 10° C., or between 10 and20° C., or between 20 and 30° C. In another embodiment is a process forthe preparation of an auris-acceptable thermoreversible gel formulationcomprising: filtering the aqueous solution containing thethermoreversible gel components at low temperature through asterilization filter; lyophilizing the sterile solution; andreconstituting the thermoreversible gel formulation with sterile waterprior to administration.

In certain embodiments, the active ingredients are dissolved in asuitable vehicle (e.g. a buffer) and sterilized separately (e.g. by heattreatment, filtration, gamma radiation). In some instances, the activeingredients are sterilized separately in a dry state. In some instances,the active ingredients are sterilized as a suspension or as a colloidalsuspension. The remaining excipients (e.g., fluid gel components presentin auris formulations) are sterilized in a separate step by a suitablemethod (e.g. filtration and/or irradiation of a cooled mixture ofexcipients); the two solutions that are separately sterilized are thenmixed aseptically to provide a final auris formulation. In someinstances, the final aseptic mixing is performed just prior toadministration of a formulation described herein.

In some instances, conventionally used methods of sterilization (e.g.,heat treatment (e.g., in an autoclave), gamma irradiation, filtration)lead to irreversible degradation of polymeric components (e.g.,thermosetting, gelling or mucoadhesive polymer components) and/or theactive agent in the formulation. In some instances, sterilization of anauris formulation by filtration through membranes (e.g., 0.2 μMmembranes) is not possible if the formulation comprises thixotropicpolymers that gel during the process of filtration.

Accordingly, provided herein are methods for sterilization of aurisformulations that prevent degradation of polymeric components (e.g.,thermosetting and/or gelling and/or mucoadhesive polymer components)and/or the active agent during the process of sterilization. In someembodiments, degradation of the active agent (e.g., any therapeutic oticagent described herein) is reduced or eliminated through the use ofspecific pH ranges for buffer components and specific proportions ofgelling agents in the formulations. In some embodiments, the choice ofan appropriate gelling agent and/or thermosetting polymer allows forsterilization of formulations described herein by filtration. In someembodiments, the use of an appropriate thermosetting polymer and anappropriate copolymer (e.g., a gelling agent) in combination with aspecific pH range for the formulation allows for high temperaturesterilization of formulations described with substantially nodegradation of the therapeutic agent or the polymeric excipients. Anadvantage of the methods of sterilization provided herein is that, incertain instances, the formulations are subjected to terminalsterilization via autoclaving without any loss of the active agentand/or excipients and/or polymeric components during the sterilizationstep and are rendered substantially free of microbes and/or pyrogens.

Microorganisms

Provided herein are auris-acceptable compositions or devices thatameliorate or lessen otic disorders described herein. Further providedherein are methods comprising the administration of said oticcompositions. In some embodiments, the compositions or devices aresubstantially free of microorganisms. Acceptable bioburden or sterilitylevels are based on applicable standards that define therapeuticallyacceptable compositions, including but not limited to United StatesPharmacopeia Chapters <1111> et seq. For example, acceptable sterility(e.g., bioburden) levels include about 10 colony forming units (cfu) pergram of formulation, about 50 cfu per gram of formulation, about 100 cfuper gram of formulation, about 500 cfu per gram of formulation or about1000 cfu per gram of formulation. In some embodiments, acceptablebioburden levels or sterility for formulations include less than 10cfu/mL, less that 50 cfu/mL, less than 500 cfu/mL or less than 1000cfu/mL microbial agents. In addition, acceptable bioburden levels orsterility include the exclusion of specified objectionablemicrobiological agents. By way of example, specified objectionablemicrobiological agents include but are not limited to Escherichia coli(E. coli), Salmonella sp., Pseudomonas aeruginosa (P. aeruginosa) and/orother specific microbial agents.

Sterility of the auris-acceptable otic therapeutic agent formulation isconfirmed through a sterility assurance program in accordance withUnited States Pharmacopeia Chapters <61>, <62> and <71>. A key componentof the sterility assurance quality control, quality assurance andvalidation process is the method of sterility testing. Sterilitytesting, by way of example only, is performed by two methods. The firstis direct inoculation wherein a sample of the composition to be testedis added to growth medium and incubated for a period of time up to 21days. Turbidity of the growth medium indicates contamination. Drawbacksto this method include the small sampling size of bulk materials whichreduces sensitivity, and detection of microorganism growth based on avisual observation. An alternative method is membrane filtrationsterility testing. In this method, a volume of product is passed througha small membrane filter paper. The filter paper is then placed intomedia to promote the growth of microorganisms. This method has theadvantage of greater sensitivity as the entire bulk product is sampled.The commercially available Millipore Steritest sterility testing systemis optionally used for determinations by membrane filtration sterilitytesting. For the filtration testing of creams or ointments Steritestfilter system No. TLHVSL210 are used. For the filtration testing ofemulsions or viscous products Steritest filter system No. TLAREM210 orTDAREM210 are used. For the filtration testing of pre-filled syringesSteritest filter system No. TTHASY210 are used. For the filtrationtesting of material dispensed as an aerosol or foam Steritest filtersystem No. TTHVA210 are used. For the filtration testing of solublepowders in ampoules or vials Steritest filter system No. TTHADA210 orTTHADV210 are used.

Testing for E. coli and Salmonella includes the use of lactose brothsincubated at 30-35° C. for 24-72 hours, incubation in MacConkey and/orEMB agars for 18-24 hours, and/or the use of Rappaport medium. Testingfor the detection of P. aeruginosa includes the use of NAC agar. UnitedStates Pharmacopeia Chapter <62> further enumerates testing proceduresfor specified objectionable microorganisms.

In certain embodiments, any controlled release formulation describedherein has less than about 60 colony forming units (CFU), less thanabout 50 colony forming units, less than about 40 colony forming units,or less than about 30 colony forming units of microbial agents per gramof formulation. In certain embodiments, the otic formulations describedherein are formulated to be isotonic with the endolymph and/or theperilymph.

Endotoxins

Provided herein are otic compositions that ameliorate or lessen oticdisorders described herein. Further provided herein are methodscomprising the administration of said otic compositions. In someembodiments, the compositions or devices are substantially free ofendotoxins. An additional aspect of the sterilization process is theremoval of by-products from the killing of microorganisms (hereinafter,“Product”). The process of depyrogenation removes pyrogens from thesample. Pyrogens are endotoxins or exotoxins which induce an immuneresponse. An example of an endotoxin is the lipopolysaccharide (LPS)molecule found in the cell wall of gram-negative bacteria. Whilesterilization procedures such as autoclaving or treatment with ethyleneoxide kill the bacteria, the LPS residue induces a proinflammatoryimmune response, such as septic shock. Because the molecular size ofendotoxins can vary widely, the presence of endotoxins is expressed in“endotoxin units” (EU). One EU is equivalent to 100 picograms of E. coliLPS. Humans can develop a response to as little as 5 EU/kg of bodyweight. The bioburden (e.g., microbial limit) and/or sterility (e.g.,endotoxin level) is expressed in any units as recognized in the art. Incertain embodiments, otic compositions described herein contain lowerendotoxin levels (e.g. <4 EU/kg of body weight of a subject) whencompared to conventionally acceptable endotoxin levels (e.g., 5 EU/kg ofbody weight of a subject). In some embodiments, the auris-acceptableotic therapeutic agent formulation has less than about 5 EU/kg of bodyweight of a subject. In other embodiments, the auris-acceptable otictherapeutic agent formulation has less than about 4 EU/kg of body weightof a subject. In additional embodiments, the auris-acceptable otictherapeutic agent formulation has less than about 3 EU/kg of body weightof a subject. In additional embodiments, the auris-acceptable otictherapeutic agent formulation has less than about 2 EU/kg of body weightof a subject.

In some embodiments, the auris-acceptable otic therapeutic agentformulation or device has less than about 5 EU/kg of formulation. Inother embodiments, the auris-acceptable otic therapeutic agentformulation has less than about 4 EU/kg of formulation. In additionalembodiments, the auris-acceptable otic therapeutic agent formulation hasless than about 3 EU/kg of formulation. In some embodiments, theauris-acceptable otic therapeutic agent formulation has less than about5 EU/kg Product. In other embodiments, the auris-acceptable otictherapeutic agent formulation has less than about 1 EU/kg Product. Inadditional embodiments, the auris-acceptable otic therapeutic agentformulation has less than about 0.2 EU/kg Product. In some embodiments,the auris-acceptable otic therapeutic agent formulation has less thanabout 5 EU/g of unit or Product. In other embodiments, theauris-acceptable otic therapeutic agent formulation has less than about4 EU/g of unit or Product. In additional embodiments, theauris-acceptable otic therapeutic agent formulation has less than about3 EU/g of unit or Product. In some embodiments, the auris-acceptableotic therapeutic agent formulation has less than about 5 EU/mg of unitor Product. In other embodiments, the auris-acceptable otic therapeuticagent formulation has less than about 4 EU/mg of unit or Product. Inadditional embodiments, the auris-acceptable otic therapeutic agentformulation has less than about 3 EU/mg of unit or Product. In certainembodiments, otic compositions described herein contain from about 1 toabout 5 EU/mL of formulation. In certain embodiments, otic compositionsdescribed herein contain from about 2 to about 5 EU/mL of formulation,from about 3 to about 5 EU/mL of formulation, or from about 4 to about 5EU/mL of formulation.

In certain embodiments, otic compositions or devices described hereincontain lower endotoxin levels (e.g. <0.5 EU/mL of formulation) whencompared to conventionally acceptable endotoxin levels (e.g., 0.5 EU/mLof formulation). In some embodiments, the auris-acceptable otictherapeutic agent formulation or device has less than about 0.5 EU/mL offormulation. In other embodiments, the auris-acceptable otic therapeuticagent formulation has less than about 0.4 EU/mL of formulation. Inadditional embodiments, the auris-acceptable otic therapeutic agentformulation has less than about 0.2 EU/mL of formulation.

Pyrogen detection, by way of example only, is performed by severalmethods. Suitable tests for sterility include tests described in UnitedStates Pharmacopoeia (USP)<71> Sterility Tests (23rd edition, 1995). Therabbit pyrogen test and the Limulus amebocyte lysate test are bothspecified in the United States Pharmacopeia Chapters <85> and <151>(USP23/NF 18, Biological Tests, The United States PharmacopeialConvention, Rockville, Md., 1995). Alternative pyrogen assays have beendeveloped based upon the monocyte activation-cytokine assay. Uniformcell lines suitable for quality control applications have been developedand have demonstrated the ability to detect pyrogenicity in samples thathave passed the rabbit pyrogen test and the Limulus amebocyte lysatetest (Taktak et al, J. Pharm. Pharmacol. (1990), 43:578-82). In anadditional embodiment, the auris-acceptable otic therapeutic agentformulation is subject to depyrogenation. In a further embodiment, theprocess for the manufacture of the auris-acceptable otic therapeuticagent formulation comprises testing the formulation for pyrogenicity. Incertain embodiments, the formulations described herein are substantiallyfree of pyrogens.

pH and Practical Osmolarity

As used herein, “practical osmolarity” means the osmolarity of aformulation that is measured by including the active agent and allexcipients except the gelling and/or the thickening agent (e.g.,polyoxyethylene-polyooxypropylene copolymers, carboxymethylcellulose orthe like). The practical osmolarity of a formulation described herein ismeasured by any suitable method, e.g., a freezing point depressionmethod as described in Viegas et. al., Int. J. Pharm., 1998, 160,157-162. In some instances, the practical osmolarity of a compositiondescribed herein is measured by vapor pressure osmometry (e.g., vaporpressure depression method) that allows for determination of theosmolarity of a composition at higher temperatures. In some instances,vapor pressure depression method allows for determination of theosmolarity of a formulation comprising a gelling agent (e.g., athermoreversible polymer) at a higher temperature wherein the gellingagent is in the form of a gel. The practical osmolality of an oticformulation described herein is from about 100 mOsm/kg to about 1000mOsm/kg, from about 200 mOsm/kg to about 800 mOsm/kg, from about 250mOsm/kg to about 500 mOsm/kg, or from about 250 mOsm/kg to about 320mOsm/kg, or from about 250 mOsm/kg to about 350 mOsm/kg or from about280 mOsm/kg to about 320 mOsm/kg. In some embodiments, the formulationsdescribed herein have a practical osmolarity of about 100 mOsm/L toabout 1000 mOsm/L, about 200 mOsm/L to about 800 mOsm/L, about 250mOsm/L to about 500 mOsm/L, about 250 mOsm/L to about 350 mOsm/L, about250 mOsm/L to about 320 mOsm/L, or about 280 mOsm/L to about 320 mOsm/L.

In some embodiments, the osmolarity at a target site of action (e.g.,the perilymph) is about the same as the delivered osmolarity (i.e.,osmolarity of materials that cross or penetrate the round windowmembrane) of any formulation described herein. In some embodiments, theformulations described herein have a deliverable osmolarity of about 150mOsm/L to about 500 mOsm/L, about 250 mOsm/L to about 500 mOsm/L, about250 mOsm/L to about 350 mOsm/L, about 280 mOsm/L to about 370 mOsm/L orabout 250 mOsm/L to about 320 mOsm/L.

The main cation present in the endolymph is potassium. In addition theendolymph has a high concentration of positively charged amino acids.The main cation present in the perilymph is sodium. In certaininstances, the ionic composition of the endolymph and perilymph regulatethe electrochemical impulses of hair cells. In certain instances, anychange in the ionic balance of the endolymph or perilymph results in aloss of hearing due to changes in the conduction of electrochemicalimpulses along otic hair cells. In some embodiments, a compositiondisclosed herein does not disrupt the ionic balance of the perilymph. Insome embodiments, a composition disclosed herein has an ionic balancethat is the same as or substantially the same as the perilymph. In someembodiments, a composition disclosed herein does not disrupt the ionicbalance of the endolymph. In some embodiments, a composition disclosedherein has an ionic balance that is the same as or substantially thesame as the endolymph. In some embodiments, an otic formulationdescribed herein is formulated to provide an ionic balance that iscompatible with inner ear fluids (e.g., endolymph and/or perilymph).

The endolymph and the perilymph have a pH that is close to thephysiological pH of blood. The endolymph has a pH range of about7.2-7.9; the perilymph has a pH range of about 7.2-7.4. The in situ pHof the proximal endolymph is about 7.4 while the pH of distal endolymphis about 7.9.

In some embodiments, the pH of a composition described herein isadjusted (e.g., by use of a buffer) to an endolymph-compatible pH rangeof about 5.5 to 9.0. In specific embodiments, the pH of a compositiondescribed herein is adjusted to a perilymph-suitable pH range of about5.5 to about 9.0. In some embodiments, the pH of a composition describedherein is adjusted to a perilymph-suitable range of about 5.5 to about8.0, about 6 to about 8.0 or about 6.6 to about 8.0. In someembodiments, the pH of a composition described herein is adjusted to aperilymph-suitable pH range of about 7.0-7.6.

In some embodiments, useful formulations also include one or more pHadjusting agents or buffering agents. Suitable pH adjusting agents orbuffers include, but are not limited to acetate, bicarbonate, ammoniumchloride, citrate, phosphate, pharmaceutically acceptable salts thereofand combinations or mixtures thereof.

In one embodiment, when one or more buffers are utilized in theformulations of the present disclosure, they are combined, e.g., with apharmaceutically acceptable vehicle and are present in the finalformulation, e.g., in an amount ranging from about 0.1% to about 20%,from about 0.5% to about 10%. In certain embodiments of the presentdisclosure, the amount of buffer included in the gel formulations are anamount such that the pH of the gel formulation does not interfere withthe body's natural buffering system.

In one embodiment, diluents are also used to stabilize compounds becausethey can provide a more stable environment. Salts dissolved in bufferedsolutions (which also can provide pH control or maintenance) areutilized as diluents in the art, including, but not limited to aphosphate buffered saline solution.

In some embodiments, any gel formulation described herein has a pH thatallows for sterilization (e.g, by filtration or aseptic mixing or heattreatment and/or autoclaving (e.g., terminal sterilization) of a gelformulation without degradation of the pharmaceutical agent (e.g., auralpressure modulating agent) or the polymers comprising the gel. In orderto reduce hydrolysis and/or degradation of the otic agent and/or the gelpolymer during sterilization, the buffer pH is designed to maintain pHof the formulation in the 7-8 range during the process of sterilization(e.g., high temperature autoclaving).

In specific embodiments, any gel formulation described herein has a pHthat allows for terminal sterilization (e.g, by heat treatment and/orautoclaving) of a gel formulation without degradation of thepharmaceutical agent (e.g., aural pressure modulating agent) or thepolymers comprising the gel. For example, in order to reduce hydrolysisand/or degradation of the otic agent and/or the gel polymer duringautoclaving, the buffer pH is designed to maintain pH of the formulationin the 7-8 range at elevated temperatures. Any appropriate buffer isused depending on the otic agent used in the formulation. In someinstances, since pK_(a) of TRIS decreases as temperature increases atapproximately −0.03/° C. and pK_(a) of PBS increases as temperatureincreases at approximately 0.003/° C., autoclaving at 250° F. (121° C.)results in a significant downward pH shift (i.e. more acidic) in theTRIS buffer whereas a relatively much less upward pH shift in the PBSbuffer and therefore much increased hydrolysis and/or degradation of anotic agent in TRIS than in PBS. Degradation of an otic agent is reducedby the use of an appropriate combination of a buffer and polymericadditives (e.g. CMC) as described herein.

In some embodiments, a formulation pH of between about 5.0 and about9.0, between about 5.5 and about 8.5, between about 6.0 and about 7.6,between about 7 and about 7.8, between about 7.0 and about 7.6, betweenabout 7.2 and 7.6, or between about 7.2 and about 7.4 is suitable forsterilization (e.g, by filtration or aseptic mixing or heat treatmentand/or autoclaving (e.g., terminal sterilization)) of auris formulationsdescribed herein. In specific embodiments a formulation pH of about 6.0,about 6.5, about 7.0, about 7.1, about 7.2, about 7.3, about 7.4, about7.5, or about 7.6 is suitable for sterilization (e.g, by filtration oraseptic mixing or heat treatment and/or autoclaving (e.g., terminalsterilization)) of any composition described herein.

In some embodiments, the formulations have a pH as described herein, andinclude a thickening agent (e.g, a viscosity enhancing agent) such as,by way of non-limiting example, a cellulose based thickening agentdescribed herein. In some instances, the addition of a secondary polymer(e.g., a thickening agent) and a pH of formulation as described herein,allows for sterilization of a formulation described herein without anysubstantial degradation of the otic agent and/or the polymer componentsin the otic formulation. In some embodiments, the ratio of athermoreversible poloxamer to a thickening agent in a formulation thathas a pH as described herein, is about 40:1, about 35:1, about 30:1,about 25:1, about 20:1, about 15:1 about 10:1, or about 5:1. Forexample, in certain embodiments, a sustained and/or extended releaseformulation described herein comprises a combination of poloxamer 407(pluronic F127) and carboxymethylcellulose (CMC) in a ratio of about40:1, about 35:1, about 30:1, about 25:1, about 20:1, about 15:1, about10:1 or about 5:1.

In some embodiments, the amount of thermoreversible polymer in anyformulation described herein is about 10%, about 15%, about 20%, about25%, about 30%, about 35% or about 40% of the total weight of theformulation. In some embodiments, the amount of thermoreversible polymerin any formulation described herein is about 10%, about 11%, about 12%,about 13%, about 14%, about 15%, about 16%, about 17%, about 18%, about19%, about 20%, about 21%, about 22%, about 23%, about 24% or about 25%of the total weight of the formulation. In some embodiments, the amountof thermoreversible polymer (e.g., pluronic F127) in any formulationdescribed herein is about 7.5% of the total weight of the formulation.In some embodiments, the amount of thermoreversible polymer (e.g.,pluronic F127) in any formulation described herein is about 10% of thetotal weight of the formulation. In some embodiments, the amount ofthermoreversible polymer (e.g., pluronic F127) in any formulationdescribed herein is about 11% of the total weight of the formulation. Insome embodiments, the amount of thermoreversible polymer (e.g., pluronicF127) in any formulation described herein is about 12% of the totalweight of the formulation. In some embodiments, the amount ofthermoreversible polymer (e.g., pluronic F127) in any formulationdescribed herein is about 13% of the total weight of the formulation. Insome embodiments, the amount of thermoreversible polymer (e.g., pluronicF127) in any formulation described herein is about 14% of the totalweight of the formulation. In some embodiments, the amount ofthermoreversible polymer (e.g., pluronic F127) in any formulationdescribed herein is about 15% of the total weight of the formulation. Insome embodiments, the amount of thermoreversible polymer (e.g., pluronicF127) in any formulation described herein is about 16% of the totalweight of the formulation. In some embodiments, the amount ofthermoreversible polymer (e.g., pluronic F127) in any formulationdescribed herein is about 17% of the total weight of the formulation. Insome embodiments, the amount of thermoreversible polymer (e.g., pluronicF127) in any formulation described herein is about 18% of the totalweight of the formulation. In some embodiments, the amount ofthermoreversible polymer (e.g., pluronic F127) in any formulationdescribed herein is about 19% of the total weight of the formulation. Insome embodiments, the amount of thermoreversible polymer (e.g., pluronicF127) in any formulation described herein is about 20% of the totalweight of the formulation. In some embodiments, the amount ofthermoreversible polymer (e.g., pluronic F127) in any formulationdescribed herein is about 21% of the total weight of the formulation. Insome embodiments, the amount of thermoreversible polymer (e.g., pluronicF127) in any formulation described herein is about 23% of the totalweight of the formulation. In some embodiments, the amount ofthermoreversible polymer (e.g., pluronic F127) in any formulationdescribed herein is about 25% of the total weight of the formulation.

In some embodiments, the amount of thickening agent (e.g., a gellingagent) in any formulation described herein is about 1%, about 5%, about10%, or about 15% of the total weight of the formulation. In someembodiments, the amount of thickening agent (e.g., a gelling agent) inany formulation described herein is about 0.5%, about 1%, about 1.5%,about 2%, about 2.5%, about 3%, about 3.5%, about 4%, about 4.5%, orabout 5% of the total weight of the formulation.

In some embodiments, the pharmaceutical formulations described hereinare stable with respect to pH over a period of any of at least about 1day, at least about 2 days, at least about 3 days, at least about 4days, at least about 5 days, at least about 6 days, at least about 1week, at least about 2 weeks, at least about 3 weeks, at least about 4weeks, at least about 5 weeks, at least about 6 weeks, at least about 7weeks, at least about 8 weeks, at least about 1 month, at least about 2months, at least about 3 months, at least about 4 months, at least about5 months, or at least about 6 months. In other embodiments, theformulations described herein are stable with respect to pH over aperiod of at least about 1 week. Also described herein are formulationsthat are stable with respect to pH over a period of at least about 1month.

Tonicity Agents

In general, the endolymph has a higher osmolality than the perilymph.For example, the endolymph has an osmolality of about 304 mOsm/kg H₂Owhile the perilymph has an osmolality of about 294 mOsm/kg H₂O. Incertain embodiments, tonicity agents are added to the formulationsdescribed herein in an amount as to provide a practical osmolality of anotic formulation of about 100 mOsm/kg to about 1000 mOsm/kg, from about200 mOsm/kg to about 800 mOsm/kg, from about 250 mOsm/kg to about 500mOsm/kg, or from about 250 mOsm/kg to about 350 mOsm/kg or from about280 mOsm/kg to about 320 mOsm/kg. In some embodiments, the formulationsdescribed herein have a practical osmolarity of about 100 mOsm/L toabout 1000 mOsm/L, about 200 mOsm/L to about 800 mOsm/L, about 250mOsm/L to about 500 mOsm/L, about 250 mOsm/L to about 350 mOsm/L, about280 mOsm/L to about 320 mOsm/L or about 250 mOsm/L to about 320 mOsm/L.

In some embodiments, the deliverable osmolarity of any formulationdescribed herein is designed to be isotonic with the targeted oticstructure (e.g., endolymph, perilymph or the like). In specificembodiments, auris compositions described herein are formulated toprovide a delivered perilymph-suitable osmolarity at the target site ofaction of about 250 to about 320 mOsm/L; and preferably about 270 toabout 320 mOsm/L. In specific embodiments, auris compositions describedherein are formulated to provide a delivered perilymph-suitableosmolality at the target site of action of about 250 to about 320mOsm/kg H₂O; or an osmolality of about 270 to about 320 mOsm/kg H₂O. Inspecific embodiments, the deliverable osmolarity/osmolality of theformulations (i.e., the osmolarity/osmolality of the formulation in theabsence of gelling or thickening agents (e.g., thermoreversible gelpolymers) is adjusted, for example, by the use of appropriate saltconcentrations (e.g., concentration of potassium or sodium salts) or theuse of tonicity agents which renders the formulationsendolymph-compatible and/or perilymph-compatible (i.e. isotonic with theendolymph and/or perilymph) upon delivery at the target site. Theosmolarity of a formulation comprising a thermoreversible gel polymer isan unreliable measure due to the association of varying amounts of waterwith the monomeric units of the polymer. The practical osmolarity of aformulation (i.e., osmolarity in the absence of a gelling or thickeningagent (e.g. a thermoreversible gel polymer) is a reliable measure and ismeasured by any suitable method (e.g., freezing point depression method,vapor depression method). In some instances, the formulations describedherein provide a deliverable osmolarity (e.g., at a target site (e.g.,perilymph) that causes minimal disturbance to the environment of theinner ear and causes minimum discomfort (e.g., vertigo and/or nausea) toa mammal upon administration.

In some embodiments, any formulation described herein is isotonic withthe perilymph and/or endolymph. Isotonic formulations are provided bythe addition of a tonicity agent. Suitable tonicity agents include, butare not limited to any pharmaceutically acceptable sugar, salt or anycombinations or mixtures thereof, such as, but not limited to dextrose,glycerin, mannitol, sorbitol, sodium chloride, and other electrolytes.

Useful auris compositions include one or more salts in an amountrequired to bring osmolality of the composition into an acceptablerange. Such salts include those having sodium, potassium or ammoniumcations and chloride, citrate, ascorbate, borate, phosphate,bicarbonate, sulfate, thiosulfate or bisulfite anions; suitable saltsinclude sodium chloride, potassium chloride, sodium thiosulfate, sodiumbisulfite and ammonium sulfate.

In some embodiments, the formulations described herein have a pH and/orpractical osmolarity as described herein, and have a concentration ofactive pharmaceutical ingredient between about 1 μM and about 10 μM,between about 1 mM and about 100 mM, between about 0.1 mM and about 100mM, between about 0.1 mM and about 100 nM. In some embodiments, theformulations described herein have a pH and/or practical osmolarity asdescribed herein, and have a concentration of active pharmaceuticalingredient between about 0.01%-about 20%, between about 0.01%-about 10%,between about 0.01%-about 7.5%, between about 0.01%-6%, between about0.01-5%, between about 0.1-about 10%, or between about 0.1-about 6% ofthe active ingredient by weight of the formulation. In some embodiments,the formulations described herein have a pH and/or practical osmolarityas described herein, and have a concentration of active pharmaceuticalingredient between about 0.1 and about 70 mg, between about 1 mg andabout 70 mg/mL, between about 1 mg and about 50 mg/mL, between about 1mg/mL and about 20 mg/mL, between about 1 mg/mL to about 10 mg/mL,between about 1 mg/mL to about 5 mg/mL, or between about 0.5 mg/mL toabout 5 mg/mL of the active agent by volume of the formulation. In someembodiments, the formulations described herein have a pH and/orpractical osmolarity as described herein, and have a concentration ofactive pharmaceutical ingredient between about 1 μg/mL and about 500μg/mL, between about 1 μg/mL and about 250 μg/mL, between about 1 μg andabout 100 μg/mL, between about 1 μg/mL and about 50 μg/mL, or betweenabout 1 μg/mL and about 20 μg/mL of the active agent by volume of theformulation.

Particle Size

Size reduction is used to increase surface area and/or modulateformulation dissolution properties. It is also used to maintain aconsistent average particle size distribution (PSD) (e.g.,micrometer-sized particles, nanometer-sized particles or the like) forany formulation described herein. In some embodiments, any formulationdescribed herein comprises multiparticulates, i.e., a plurality ofparticle sizes (e.g., micronized particles, nano-sized particles,non-sized particles, colloidal particles); i.e, the formulation is amultiparticulate formulation. In some embodiments, any formulationdescribed herein comprises one or more multiparticulate (e.g.,micronized) therapeutic agents. Micronization is a process of reducingthe average diameter of particles of a solid material. Micronizedparticles are from about micrometer-sized in diameter to aboutnanometer-sized in diameter. In some embodiments, the average diameterof particles in a micronized solid is from about 0.5 μm to about 500 μm.In some embodiments, the average diameter of particles in a micronizedsolid is from about 1 μm to about 200 μm. In some embodiments, theaverage diameter of particles in a micronized solid is from about 2 μmto about 100 μm. In some embodiments, the average diameter of particlesin a micronized solid is from about 3 μm to about 50 μm. In someembodiments, a particulate micronized solid comprises particle sizes ofless than about 5 microns, less than about 20 microns and/or less thanabout 100 microns. In some embodiments, the use of particulates (e.g.,micronized particles) of aural pressure modulating agent allows forextended and/or sustained release of the aural pressure modulating agentfrom any formulation described herein compared to a formulationcomprising non-multiparticulate (e.g, non-micronized) aural pressuremodulating agent. In some instances, formulations containingmultiparticulate (e.g. micronized) aural pressure modulating agent areejected from a 1 mL syringe adapted with a 27 G needle without anyplugging or clogging.

In some instances, any particle in any formulation described herein is acoated particle (e.g., a coated micronized particle, nano-particle)and/or a microsphere and/or a liposomal particle. Particle sizereduction techniques include, by way of example, grinding, milling(e.g., air-attrition milling (jet milling), ball milling), coacervation,complex coacervation, high pressure homogenization, spray drying and/orsupercritical fluid crystallization. In some instances, particles aresized by mechanical impact (e.g., by hammer mills, ball mill and/or pinmills). In some instances, particles are sized via fluid energy (e.g.,by spiral jet mills, loop jet mills, and/or fluidized bed jet mills). Insome embodiments formulations described herein comprise crystallineparticles and/or isotropic particles. In some embodiments, formulationsdescribed herein comprise amorphous particles and/or anisotropicparticles. In some embodiments, formulations described herein comprisetherapeutic agent particles wherein the therapeutic agent is a freebase, or a salt, or a prodrug of a therapeutic agent, or any combinationthereof.

In some embodiments, a formulation described herein comprises one ormore aural pressure modulating agents wherein the aural pressuremodulating agent comprises nanoparticulates. In some embodiments, aformulation described herein comprises aural pressure modulating agentbeads (e.g., conivaptan beads) that are optionally coated withcontrolled release excipients. In some embodiments, a formulationdescribed herein comprises a aural pressure modulating agent that isgranulated and/or reduced in size and coated with controlled releaseexcipients; the granulated coated aural pressure modulating agentparticulates are then optionally micronized and/or formulated in any ofthe compositions described herein.

In some instances, a combination of an aural pressure modulating agentas a neutral molecule, free acid or free base and a salt of the auralpressure modulating agent is used to prepare pulsed release otic agentformulations using the procedures described herein. In someformulations, a combination of a micronized aural pressure modulatingagent (and/or salt or prodrug thereof) and coated particles (e.g.,nanoparticles, liposomes, microspheres) is used to prepare pulsedrelease otic agent formulations using any procedure described herein.Alternatively, a pulsed release profile is achieved by solubilizing upto 20% of the delivered dose of the aural pressure modulating agent(e.g., micronized aural pressure modulating agent, free base, free acidor salt or prodrug thereof; multiparticulate aural pressure modulatingagent, free base, free acid or salt or prodrug thereof) with the aid ofcyclodextrins, surfactants (e.g., poloxamers (407, 338, 188), tween (80,60, 20, 81), PEG-hydrogenated castor oil, cosolvents likeN-methyl-2-Pyrrolidone or the like and preparing pulsed releaseformulations using any procedure described herein.

In specific embodiments, any auris-compatible formulation describedherein comprises one or more micronized pharmaceutical agents (e.g.,aural pressure modulating agents). In some of such embodiments, amicronized pharmaceutical agent comprises micronized particles, coated(e.g., with an extended release coat) micronized particles, or acombination thereof. In some of such embodiments, a micronizedpharmaceutical agent comprising micronized particles, coated micronizedparticles, or a combination thereof, comprises an aural pressuremodulating agent as a neutral molecule, a free acid, a free base, asalt, a prodrug or any combination thereof. In certain embodiments, apharmaceutical composition described herein comprises an aural pressuremodulating agent as a micronized powder. In certain embodiments, apharmaceutical composition described herein comprises an aural pressuremodulating agent in the form of a micro-aural pressure modulating agentpowder.

The multiparticulates and/or micronized aural pressure modulating agentsdescribed herein are delivered to an auris structure (e.g., inner ear)by means of any type of matrix including solid, liquid or gel matrices.In some embodiments, the multiparticulates and/or micronized auralpressure modulating agents described herein are delivered to an aurisstructure (e.g., inner ear) by means of any type of matrix includingsolid, liquid or gel matrices via intratympanic injection.

Pharmaceutical Formulations

Provided herein are pharmaceutical compositions or devices that includeat least one aural pressure modulating agent and a pharmaceuticallyacceptable diluent(s), excipient(s), or carrier(s). In some embodiments,the pharmaceutical compositions include other medicinal orpharmaceutical agents, carriers, adjuvants, such as preserving,stabilizing, wetting or emulsifying agents, solution promoters, saltsfor regulating the osmotic pressure, and/or buffers. In otherembodiments, the pharmaceutical compositions also contain othertherapeutic substances.

In some embodiments, the compositions or devices described hereininclude a dye to help enhance the visualization of the gel when applied.In some embodiments, dyes that are compatible with the auris-acceptablecompositions or devices described herein include Evans blue (e.g., 0.5%of the total weight of an otic formulation), Methylene blue (e.g., 1% ofthe total weight of an otic formulation), Isosulfan blue (e.g., 1% ofthe total weight of an otic formulation), Trypan blue (e.g., 0.15% ofthe total weight of an otic formulation), and/or indocyanine green(e.g., 25 mg/vial). Other common dyes, e.g, FD&C red 40, FD&C red 3,FD&C yellow 5, FD&C yellow 6, FD&C blue 1, FD&C blue2, FD&C green 3,fluorescence dyes (e.g., Fluorescein isothiocyanate, rhodamine, AlexaFluors, DyLight Fluors) and/or dyes that are visualizable in conjunctionwith non-invasive imaging techniques such as MRI, CAT scans, PET scansor the like. Gadolinium-based MRI dyes, iodine-base dyes, barium-baseddyes or the like are also contemplated for use with any otic formulationdescribed herein. Other dyes that are compatible with any formulation orcomposition described herein are listed in the Sigma-Aldrich catalogunder dyes (which is included herein by reference for such disclosure).

Any pharmaceutical composition or device described herein isadministered by locating the composition or device in contact with thecrista fenestrae cochlea, the round window, the tympanic cavity, thetympanic membrane, the auris media or the auris externa.

In one specific embodiment of the auris-acceptable controlled releaseaural pressure modulating agent pharmaceutical formulations describedherein, the aural pressure modulating agent is provided in a gel matrix,also referred to herein as “auris acceptable gel formulations,” “aurisinterna-acceptable gel formulations,” “auris media-acceptable gelformulations,” “auris externa-acceptable gel formulations”, “auris gelformulations” or variations thereof. All of the components of the gelformulation must be compatible with the targeted auris structure.Further, the gel formulations provide controlled release of the auralpressure modulating agent to the desired site within the targeted aurisstructure; in some embodiments, the gel formulation also has animmediate or rapid release component for delivery of the aural pressuremodulating agent to the desired target site. In other embodiments, thegel formulation has a sustained release component for delivery of theaural pressure modulating agent. In some embodiments, the gelformulation comprises a multiparticulate (e.g., micronized) auralpressure modulating agent. In some embodiments, the auris gelformulations are biodegradeable. In other embodiments, the auris gelformulations include a mucoadhesive excipient to allow adhesion to theexternal mucous layer of the round window membrane. In yet otherembodiments, the auris gel formulations include a penetration enhancerexcipient; in further embodiments, the auris gel formulation contains aviscosity enhancing agent sufficient to provide a viscosity of betweenabout 500 and 1,000,000 centipoise, between about 750 and 1,000,000centipoise; between about 1000 and 1,000,000 centipoise; between about1000 and 400,000 centipoise; between about 2000 and 100,000 centipoise;between about 3000 and 50,000 centipoise; between about 4000 and 25,000centipoise; between about 5000 and 20,000 centipoise; or between about6000 and 15,000 centipoise. In some embodiments, the auris gelformulation contains a viscosity enhancing agent sufficient to provide aviscosity of between about 50,0000 and 1,000,000 centipoise.

In other embodiments, the auris interna pharmaceutical formulationsdescribed herein further provide an auris-acceptable hydrogel; in yetother embodiments, the auris pharmaceutical formulations provide anauris-acceptable microsphere or microparticle; in still otherembodiments, the auris pharmaceutical formulations provide anauris-acceptable liposome. In some embodiments, the auris pharmaceuticalformulations provide an auris-acceptable foam; in yet other embodiments,the auris pharmaceutical formulations provide an auris-acceptable paint;in still further embodiments, the auris pharmaceutical formulationsprovide an auris-acceptable in situ forming spongy material. In someembodiments, the auris pharmaceutical formulations provide anauris-acceptable solvent release gel. In some embodiments, the aurispharmaceutical formulations provide an actinic radiation curable gel.Further embodiments include a thermoreversible gel in the aurispharmaceutical formulation, such that upon preparation of the gel atroom temperature or below, the formulation is a fluid, but uponapplication of the gel into or near the auris interna and/or auris mediatarget site, including the tympanic cavity, round window membrane or thecrista fenestrae cochleae, the auris-pharmaceutical formulation stiffensor hardens into a gel-like substance.

In further or alternative embodiments, the auris gel formulations arecapable of being administered on or near the round window membrane viaintratympanic injection. In other embodiments, the auris gelformulations are administered on or near the round window or the cristafenestrae cochleae through entry via a post-auricular incision andsurgical manipulation into or near the round window or the cristafenestrae cochleae area. Alternatively, the auris gel formulation isapplied via syringe and needle, wherein the needle is inserted throughthe tympanic membrane and guided to the area of the round window orcrista fenestrae cochleae. The auris gel formulations are then depositedon or near the round window or crista fenestrae cochleae for localizedtreatment of autoimmune otic disorders. In other embodiments, the aurisgel formulations are applied via microcathethers implanted into thepatient, and in yet further embodiments the formulations areadministered via a pump device onto or near the round window membrane.In still further embodiments, the auris gel formulations are applied ator near the round window membrane via a microinjection device. In yetother embodiments, the auris gel formulations are applied in thetympanic cavity. In some embodiments, the auris gel formulations areapplied on the tympanic membrane. In still other embodiments, the aurisgel formulations are applied onto or in the auditory canal.

In further specific embodiments, any pharmaceutical composition ordevice described herein comprises a multiparticulate aural pressuremodulating agentin a liquid matrix (e.g., a liquid composition forintratympanic injection, or otic drops). In certain embodiments, anypharmaceutical composition described herein comprises a multiparticulateaural pressure modulating agentin a solid matrix.

Controlled Release Formulations

In general, controlled release drug formulations impart control over therelease of drug with respect to site of release and time of releasewithin the body. As discussed herein, controlled release refers toimmediate release, delayed release, sustained release, extended release,variable release, pulsatile release and bi-modal release. Manyadvantages are offered by controlled release. First, controlled releaseof a pharmaceutical agent allows less frequent dosing and thus minimizesrepeated treatment. Second, controlled release treatment results in moreefficient drug utilization and less of the compound remains as aresidue. Third, controlled release offers the possibility of localizeddrug delivery by placement of a delivery device or formulation at thesite of disease. Still further, controlled release offers theopportunity to administer and release two or more different drugs, eachhaving a unique release profile, or to release the same drug atdifferent rates or for different durations, by means of a single dosageunit.

Accordingly, one aspect of the embodiments disclosed herein is toprovide a controlled release aural pressure modulating agentauris-acceptable composition or device for the treatment of autoimmunedisorders and/or inflammatory disorders. The controlled release aspectof the compositions and/or formulations and/or devices disclosed hereinis imparted through a variety of agents, including but not limited toexcipients, agents or materials that are acceptable for use in the aurisinterna or other otic structure. By way of example only, suchexcipients, agents or materials include an auris-acceptable polymer, anauris-acceptable viscosity enhancing agent, an auris-acceptable gel, anauris-acceptable paint, an auris-acceptable foam, an auris-acceptablexerogel, an auris-acceptable microsphere or microparticle, anauris-acceptable hydrogel, an auris-acceptable in situ forming spongymaterial, an auris-acceptable actinic radiation curable gel, anauris-acceptable solvent release gel, an auris-acceptable liposome, anauris-acceptable nanocapsule or nano sphere, an auris-acceptablethermoreversible gel, or combinations thereof.

Auris-Acceptable Gels

Gels, sometimes referred to as jellies, have been defined in variousways. For example, the United States Pharmacopoeia defines gels assemisolid systems consisting of either suspensions made up of smallinorganic particles or large organic molecules interpenetrated by aliquid. Gels include a single-phase or a two-phase system. Asingle-phase gel consists of organic macromolecules distributeduniformly throughout a liquid in such a manner that no apparentboundaries exist between the dispersed macromolecules and the liquid.Some single-phase gels are prepared from synthetic macromolecules (e.g.,carbomer) or from natural gums, (e.g., tragacanth). In some embodiments,single-phase gels are generally aqueous, but will also be made usingalcohols and oils. Two-phase gels consist of a network of small discreteparticles.

Gels can also be classified as being hydrophobic or hydrophilic. Incertain embodiments, the base of a hydrophobic gel consists of a liquidparaffin with polyethylene or fatty oils gelled with colloidal silica,or aluminum or zinc soaps. In contrast, the base of hydrophobic gelsusually consists of water, glycerol, or propylene glycol gelled with asuitable gelling agent (e.g., tragacanth, starch, cellulose derivatives,carboxyvinylpolymers, and magnesium-aluminum silicates). In certainembodiments, the rheology of the compositions or devices disclosedherein is pseudo plastic, plastic, thixotropic, or dilatant.

In one embodiment the enhanced viscosity auris-acceptable formulationdescribed herein is not a liquid at room temperature. In certainembodiments, the enhanced viscosity formulation is characterized by aphase transition between room temperature and body temperature(including an individual with a serious fever, e.g., up to about 42°C.). In some embodiments, the phase transition occurs at 1° C. belowbody temperature, at 2° C. below body temperature, at 3° C. below bodytemperature, at 4° C. below body temperature, at 6° C. below bodytemperature, at 8° C. below body temperature, or at 10° C. below bodytemperature. In some embodiments, the phase transition occurs at about15° C. below body temperature, at about 20° C. below body temperature orat about 25° C. below body temperature. In specific embodiments, thegelation temperature (Tgel) of a formulation described herein is about20° C., about 25° C., or about 30° C. In certain embodiments, thegelation temperature (Tgel) of a formulation described herein is about35° C., or about 40° C. In one embodiment, administration of anyformulation described herein at about body temperature reduces orinhibits vertigo associated with intratympanic administration of oticformulations. Included within the definition of body temperature is thebody temperature of a healthy individual, or an unhealthy individual,including an individual with a fever (up to ˜42° C.). In someembodiments, the pharmaceutical compositions or devices described hereinare liquids at about room temperature and are administered at or aboutroom temperature, reducing or ameliorating side effects such as, forexample, vertigo.

Polymers composed of polyoxypropylene and polyoxyethylene formthermoreversible gels when incorporated into aqueous solutions. Thesepolymers have the ability to change from the liquid state to the gelstate at temperatures close to body temperature, therefore allowinguseful formulations that are applied to the targeted auris structure(s).The liquid state-to-gel state phase transition is dependent on thepolymer concentration and the ingredients in the solution.

Poloxamer 407 (PF-127) is a nonionic surfactant composed ofpolyoxyethylene-polyoxypropylene copolymers. Other poloxamers include188 (F-68 grade), 237 (F-87 grade), 338 (F-108 grade). Aqueous solutionsof poloxamers are stable in the presence of acids, alkalis, and metalions. PF-127 is a commercially availablepolyoxyethylene-polyoxypropylene triblock copolymer of general formulaE106 P70 E106, with an average molar mass of 13,000. The polymer can befurther purified by suitable methods that will enhance gelationproperties of the polymer. It contains approximately 70% ethylene oxide,which accounts for its hydrophilicity. It is one of the series ofpoloxamer ABA block copolymers, whose members share the chemical formulashown below.

PF-127 is of particular interest since concentrated solutions (>20% w/w)of the copolymer are transformed from low viscosity transparentsolutions to solid gels on heating to body temperature. This phenomenon,therefore, suggests that when placed in contact with the body, the gelpreparation will form a semi-solid structure and a sustained releasedepot. Furthermore, PF-127 has good solubilizing capacity, low toxicityand is, therefore, considered a good medium for drug delivery systems.

In an alternative embodiment, the thermogel is a PEG-PLGA-PEG triblockcopolymer (Jeong et al, Nature (1997), 388:860-2; Jeong et al, J.Control. Release (2000), 63:155-63; Jeong et al, Adv. Drug Delivery Rev.(2002), 54:37-51). The polymer exhibits sol-gel behavior over aconcentration of about 5% w/w to about 40% w/w. Depending on theproperties desired, the lactide/glycolide molar ratio in the PLGAcopolymer ranges from about 1:1 to about 20:1. The resulting copolymersare soluble in water and form a free-flowing liquid at room temperature,but form a hydrogel at body temperature. A commercially availablePEG-PLGA-PEG triblock copolymer is RESOMER RGP t50106 manufactured byBoehringer Ingelheim. This material is composed of a PGLA copolymer of50:50 poly(DL-lactide-co-glycolide) and is 10% w/w of PEG and has amolecular weight of about 6000.

ReGel® is a tradename of MacroMed Incorporated for a class of lowmolecular weight, biodegradable block copolymers having reverse thermalgelation properties as described in U.S. Pat. Nos. 6,004,573, 6,117,949,6,201,072, and 6,287,588. It also includes biodegradable polymeric drugcarriers disclosed in pending U.S. patent application Ser. Nos.09/906,041, 09/559,799 and 10/919,603. The biodegradable drug carriercomprises ABA-type or BAB-type triblock copolymers or mixtures thereof,wherein the A-blocks are relatively hydrophobic and comprisebiodegradable polyesters or poly(orthoester)s, and the B-blocks arerelatively hydrophilic and comprise polyethylene glycol (PEG), saidcopolymers having a hydrophobic content of between 50.1 to 83% by weightand a hydrophilic content of between 17 to 49.9% by weight, and anoverall block copolymer molecular weight of between 2000 and 8000Daltons. The drug carriers exhibit water solubility at temperaturesbelow normal mammalian body temperatures and undergo reversible thermalgelation to then exist as a gel at temperatures equal to physiologicalmammalian body temperatures. The biodegradable, hydrophobic A polymerblock comprises a polyester or poly(ortho ester), in which the polyesteris synthesized from monomers selected from the group consisting ofD,L-lactide, D-lactide, L-lactide, D,L-lactic acid, D-lactic acid,L-lactic acid, glycolide, glycolic acid, ε-caprolactone,ε-hydroxyhexanoic acid, γ-butyrolactone, γ-hydroxybutyric acid,δ-valerolactone, δ-hydroxyvaleric acid, hydroxybutyric acids, malicacid, and copolymers thereof and having an average molecular weight ofbetween about 600 and 3000 Daltons. The hydrophilic B-block segment ispreferably polyethylene glycol (PEG) having an average molecular weightof between about 500 and 2200 Daltons.

Additional biodegradable thermoplastic polyesters include AtriGel®(provided by Atrix Laboratories, Inc.) and/or those disclosed, e.g., inU.S. Pat. Nos. 5,324,519; 4,938,763; 5,702,716; 5,744,153; and5,990,194; wherein the suitable biodegradable thermoplastic polyester isdisclosed as a thermoplastic polymer. Examples of suitable biodegradablethermoplastic polyesters include polylactides, polyglycolides,polycaprolactones, copolymers thereof, terpolymers thereof, and anycombinations thereof. In some such embodiments, the suitablebiodegradable thermoplastic polyester is a polylactide, a polyglycolide,a copolymer thereof, a terpolymer thereof, or a combination thereof. Inone embodiment, the biodegradable thermoplastic polyester is 50/50poly(DL-lactide-co-glycolide) having a carboxy terminal group; ispresent in about 30 wt. % to about 40 wt. % of the composition; and hasan average molecular weight of about 23,000 to about 45,000.Alternatively, in another embodiment, the biodegradable thermoplasticpolyester is 75/25 poly(DL-lactide-co-glycolide) without a carboxyterminal group; is present in about 40 wt. % to about 50 wt. % of thecomposition; and has an average molecular weight of about 15,000 toabout 24,000. In further or alternative embodiments, the terminal groupsof the poly(DL-lactide-co-glycolide) are either hydroxyl, carboxyl, orester depending upon the method of polymerization. Polycondensation oflactic or glycolic acid provides a polymer with terminal hydroxyl andcarboxyl groups. Ring-opening polymerization of the cyclic lactide orglycolide monomers with water, lactic acid, or glycolic acid providespolymers with the same terminal groups. However, ring-opening of thecyclic monomers with a monofunctional alcohol such as methanol, ethanol,or 1-dodecanol provides a polymer with one hydroxyl group and one esterterminal groups. Ring-opening polymerization of the cyclic monomers witha diol such as 1,6-hexanediol or polyethylene glycol provides a polymerwith only hydroxyl terminal groups.

Since the polymer systems of thermoreversible gels dissolve morecompletely at reduced temperatures, methods of solubilization includeadding the required amount of polymer to the amount of water to be usedat reduced temperatures. Generally after wetting the polymer by shaking,the mixture is capped and placed in a cold chamber or in a thermostaticcontainer at about 0-10° C. in order to dissolve the polymer. Themixture is stirred or shaken to bring about a more rapid dissolution ofthe thermoreversible gel polymer. The aural pressure modulating agentand various additives such as buffers, salts, and preservatives aresubsequently added and dissolved. In some instances the aural pressuremodulating agent and/or other pharmaceutically active agent is suspendedif it is insoluble in water. The pH is modulated by the addition ofappropriate buffering agents. round window membrane mucoadhesivecharacteristics are optionally imparted to a thermoreversible gel byincorporation of round window membrane mucoadhesive carbomers, such asCarbopol® 934P, to the composition (Majithiya et al, AAPS PharmSciTech(2006), 7 (3), p. E1; EP0551626, both of which is incorporated herein byreference for such disclosure).

In one embodiment are auris-acceptable pharmaceutical gel formulationswhich do not require the use of an added viscosity enhancing agent. Suchgel formulations incorporate at least one pharmaceutically acceptablebuffer. In one aspect is a gel formulation comprising an aural pressuremodulating agent and a pharmaceutically acceptable buffer. In anotherembodiment, the pharmaceutically acceptable excipient or carrier is agelling agent.

In other embodiments, useful aural pressure modulating agentauris-acceptable pharmaceutical formulations also include one or more pHadjusting agents or buffering agents to provide an endolymph orperilymph suitable pH. Suitable pH adjusting agents or buffers include,but are not limited to acetate, bicarbonate, ammonium chloride, citrate,phosphate, pharmaceutically acceptable salts thereof and combinations ormixtures thereof. Such pH adjusting agents and buffers are included inan amount required to maintain pH of the composition between a pH ofabout 5 and about 9, in one embodiment a pH between about 6.5 to about7.5, and in yet another embodiment at a pH of about 6.5, 6.6, 6.7, 6.8,6.9, 7.0, 7.1, 7.2, 7.3, 7.4, 7.5. In one embodiment, when one or morebuffers are utilized in the formulations of the present disclosure, theyare combined, e.g., with a pharmaceutically acceptable vehicle and arepresent in the final formulation, e.g., in an amount ranging from about0.1% to about 20%, from about 0.5% to about 10%. In certain embodimentsof the present disclosure, the amount of buffer included in the gelformulations are an amount such that the pH of the gel formulation doesnot interfere with the auris media or auris interna's natural bufferingsystem, or does not interfere with the natural pH of the endolymph orperilymph: depending on where in the cochlea the aural pressuremodulating agent formulation is targeted. In some embodiments, fromabout 10 μM to about 200 mM concentration of a buffer is present in thegel formulation. In certain embodiments, from about a 5 mM to about a200 mM concentration of a buffer is present. In certain embodiments,from about a 20 mM to about a 100 mM concentration of a buffer ispresent. In one embodiment is a buffer such as acetate or citrate atslightly acidic pH. In one embodiment the buffer is a sodium acetatebuffer having a pH of about 4.5 to about 6.5. In one embodiment thebuffer is a sodium citrate buffer having a pH of about 5.0 to about 8.0,or about 5.5 to about 7.0.

In an alternative embodiment, the buffer used istris(hydroxymethyl)aminomethane, bicarbonate, carbonate or phosphate atslightly basic pH. In one embodiment, the buffer is a sodium bicarbonatebuffer having a pH of about 6.5 to about 8.5, or about 7.0 to about 8.0.In another embodiment the buffer is a sodium phosphate dibasic bufferhaving a pH of about 6.0 to about 9.0.

Also described herein are controlled release formulations or devicescomprising a aural pressure modulating agent and a viscosity enhancingagent. Suitable viscosity-enhancing agents include by way of exampleonly, gelling agents and suspending agents. In one embodiment, theenhanced viscosity formulation does not include a buffer. In otherembodiments, the enhanced viscosity formulation includes apharmaceutically acceptable buffer. Sodium chloride or other tonicityagents are optionally used to adjust tonicity, if necessary.

By way of example only, the auris-acceptable viscosity agent includehydroxypropyl methylcellulose, hydroxyethyl cellulose,polyvinylpyrrolidone, carboxymethyl cellulose, polyvinyl alcohol, sodiumchondroitin sulfate, sodium hyaluronate. Other viscosity enhancingagents compatible with the targeted auris structure include, but are notlimited to, acacia (gum arabic), agar, aluminum magnesium silicate,sodium alginate, sodium stearate, bladderwrack, bentonite, carbomer,carrageenan, Carbopol, xanthan, cellulose, microcrystalline cellulose(MCC), ceratonia, chitin, carboxymethylated chitosan, chondrus,dextrose, furcellaran, gelatin, Ghatti gum, guar gum, hectorite,lactose, sucrose, maltodextrin, mannitol, sorbitol, honey, maize starch,wheat starch, rice starch, potato starch, gelatin, sterculia gum,xanthum gum, gum tragacanth, ethyl cellulose, ethylhydroxyethylcellulose, ethylmethyl cellulose, methyl cellulose, hydroxyethylcellulose, hydroxyethylmethyl cellulose, hydroxypropyl cellulose,poly(hydroxyethyl methacrylate), oxypolygelatin, pectin, polygeline,povidone, propylene carbonate, methyl vinyl ether/maleic anhydridecopolymer (PVM/MA), poly(methoxyethyl methacrylate),poly(methoxyethoxyethyl methacrylate), hydroxypropyl cellulose,hydroxypropylmethyl-cellulose (HPMC), sodium carboxymethyl-cellulose(CMC), silicon dioxide, polyvinylpyrrolidone (PVP: povidone), Splenda®(dextrose, maltodextrin and sucralose) or combinations thereof. Inspecific embodiments, the viscosity-enhancing excipient is a combinationof MCC and CMC. In another embodiment, the viscosity-enhancing agent isa combination of carboxymethylated chitosan, or chitin, and alginate.The combination of chitin and alginate with the aural pressuremodulating agents disclosed herein acts as a controlled releaseformulation, restricting the diffusion of the aural pressure modulatingagents from the formulation. Moreover, the combination ofcarboxymethylated chitosan and alginate is optionally used to assist inincreasing the permeability of the aural pressure modulating agentsthrough the round window membrane.

In some embodiments is an enhanced viscosity formulation, comprisingfrom about 0.1 mM and about 100 mM of a aural pressure modulating agent,a pharmaceutically acceptable viscosity agent, and water for injection,the concentration of the viscosity agent in the water being sufficientto provide a enhanced viscosity formulation with a final viscosity fromabout 100 to about 100,000 cP. In certain embodiments, the viscosity ofthe gel is in the range from about 100 to about 50,000 cP, about 100 cPto about 1,000 cP, about 500 cP to about 1500 cP, about 1000 cP to about3000 cP, about 2000 cP to about 8,000 cP, about 4,000 cP to about 50,000cP, about 10,000 cP to about 500,000 cP, about 15,000 cP to about1,000,000 cP. In other embodiments, when an even more viscous medium isdesired, the biocompatible gel comprises at least about 35%, at leastabout 45%, at least about 55%, at least about 65%, at least about 70%,at least about 75%, or even at least about 80% or so by weight of theaural pressure modulating agent. In highly concentrated samples, thebiocompatible enhanced viscosity formulation comprises at least about25%, at least about 35%, at least about 45%, at least about 55%, atleast about 65%, at least about 75%, at least about 85%, at least about90% or at least about 95% or more by weight of the aural pressuremodulating agent.

In some embodiments, the viscosity of the gel formulations presentedherein are measured by any means described. For example, in someembodiments, an LVDV-II+CP Cone Plate Viscometer and a Cone SpindleCPE-40 is used to calculate the viscosity of the gel formulationdescribed herein. In other embodiments, a Brookfield (spindle and cup)viscometer is used to calculate the viscosity of the gel formulationdescribed herein. In some embodiments, the viscosity ranges referred toherein are measured at room temperature. In other embodiments, theviscosity ranges referred to herein are measured at body temperature(e.g., at the average body temperature of a healthy human).

In one embodiment, the pharmaceutically acceptable enhanced viscosityauris-acceptable formulation comprises at least one aural pressuremodulating agent and at least one gelling agent. Suitable gelling agentsfor use in preparation of the gel formulation include, but are notlimited to, celluloses, cellulose derivatives, cellulose ethers (e.g.,carboxymethylcellulose, ethylcellulose, hydroxyethylcellulose,hydroxymethylcellulose, hydroxypropylmethylcellulose,hydroxypropylcellulose, methylcellulose), guar gum, xanthan gum, locustbean gum, alginates (e.g., alginic acid), silicates, starch, tragacanth,carboxyvinyl polymers, carrageenan, paraffin, petrolatum and anycombinations or mixtures thereof. In some other embodiments,hydroxypropylmethylcellulose (Methocel®) is utilized as the gellingagent. In certain embodiments, the viscosity enhancing agents describedherein are also utilized as the gelling agent for the gel formulationspresented herein.

In some embodiments, the otic therapeutic agents disclosed herein aredispensed as an auris-acceptable paint. As used herein, paints (alsoknown as film formers) are solutions comprised of a solvent, a monomeror polymer, an active agent, and optionally one or morepharmaceutically-acceptable excipients. After application to a tissue,the solvent evaporates leaving behind a thin coating comprised of themonomers or polymers, and the active agent. The coating protects activeagents and maintains them in an immobilized state at the site ofapplication. This decreases the amount of active agent which may be lostand correspondingly increases the amount delivered to the subject. Byway of non-limiting example, paints include collodions (e.g. FlexibleCollodion, USP), and solutions comprising saccharide siloxane copolymersand a cross-linking agent. Collodions are ethyl ether/ethanol solutionscontaining pyroxylin (a nitrocellulose). After application, the ethylether/ethanol solution evaporates leaving behind a thin film ofpyroxylin. In solutions comprising saccharide siloxane copolymers, thesaccharide siloxane copolymers form the coating after evaporation of thesolvent initiates the cross-linking of the saccharide siloxanecopolymers. For additional disclosures regarding paints, see Remington:The Science and Practice of Pharmacy which is hereby incorporated withrespect to this subject matter. The paints contemplated for use herein,are flexible such that they do not interfere with the propagation ofpressure waves through the ear. Further, the paints may be applied as aliquid (i.e. solution, suspension, or emulsion), a semisolid (i.e. agel, foam, paste, or jelly) or an aerosol.

In some embodiments, the otic therapeutic agents disclosed herein aredispensed as a controlled-release foam. Examples of suitable foamablecarriers for use in the compositions disclosed herein include, but arenot limited to, alginate and derivatives thereof, carboxymethylcelluloseand derivatives thereof, collagen, polysaccharides, including, forexample, dextran, dextran derivatives, pectin, starch, modified starchessuch as starches having additional carboxyl and/or carboxamide groupsand/or having hydrophilic side-chains, cellulose and derivativesthereof, agar and derivatives thereof, such as agar stabilised withpolyacrylamide, polyethylene oxides, glycol methacrylates, gelatin, gumssuch as xanthum, guar, karaya, gellan, arabic, tragacanth and locustbean gum, or combinations thereof. Also suitable are the salts of theaforementioned carriers, for example, sodium alginate. The formulationoptionally further comprises a foaming agent, which promotes theformation of the foam, including a surfactant or external propellant.Examples of suitable foaming agents include cetrimide, lecithin, soaps,silicones and the like. Commercially available surfactants such asTween® are also suitable.

In some embodiments, other gel formulations are useful depending uponthe particular aural pressure modulating agent, other pharmaceuticalagent or excipients/additives used, and as such are considered to fallwithin the scope of the present disclosure. For example, othercommercially-available glycerin-based gels, glycerin-derived compounds,conjugated, or crosslinked gels, matrices, hydrogels, and polymers, aswell as gelatins and their derivatives, alginates, and alginate-basedgels, and even various native and synthetic hydrogel andhydrogel-derived compounds are all expected to be useful in the auralpressure modulating agent formulations described herein. In someembodiments, auris-acceptable gels include, but are not limited to,alginate hydrogels SAF®-Gel (ConvaTec, Princeton, N.J.), Duoderm®Hydroactive Gel (ConvaTec), Nu-gel® (Johnson & Johnson Medical,Arlington, Tex.); Carrasyn® (V) Acemannan Hydrogel (CarringtonLaboratories, Inc., Irving, Tex.); glycerin gels Elta® Hydrogel(Swiss-American Products, Inc., Dallas, Tex.) and K-Y® Sterile (Johnson& Johnson). In further embodiments, biodegradable biocompatible gelsalso represent compounds present in auris-acceptable formulationsdisclosed and described herein.

In some formulations developed for administration to a mammal, and forcompositions formulated for human administration, the auris-acceptablegel comprises substantially all of the weight of the composition. Inother embodiments, the auris-acceptable gel comprises as much as about98% or about 99% of the composition by weight. This is desirous when asubstantially non-fluid, or substantially viscous formulation is needed.In a further embodiment, when slightly less viscous, or slightly morefluid auris-acceptable pharmaceutical gel formulations are desired, thebiocompatible gel portion of the formulation comprises at least about50% by weight, at least about 60% by weight, at least about 70% byweight, or even at least about 80% or 90% by weight of the compound. Allintermediate integers within these ranges are contemplated to fallwithin the scope of this disclosure, and in some alternativeembodiments, even more fluid (and consequently less viscous)auris-acceptable gel compositions are formulated, such as for example,those in which the gel or matrix component of the mixture comprises notmore than about 50% by weight, not more than about 40% by weight, notmore than about 30% by weight, or even those than comprise not more thanabout 15% or about 20% by weight of the composition.

Auris-Acceptable Suspending Agents

In one embodiment, at least one aural pressure modulating agent isincluded in a pharmaceutically acceptable enhanced viscosity formulationwherein the formulation further comprises at least one suspending agent,wherein the suspending agent assists in imparting controlled releasecharacteristics to the formulation. In some embodiments, suspendingagents also serve to increase the viscosity of the auris-acceptableaural pressure modulating formulations and compositions.

Suspending agents include, by way of example only, compounds such aspolyvinylpyrrolidone, e.g., polyvinylpyrrolidone K12,polyvinylpyrrolidone K17, polyvinylpyrrolidone K25, orpolyvinylpyrrolidone K30, vinyl pyrrolidone/vinyl acetate copolymer(S630), sodium carboxymethylcellulose, methylcellulose,hydroxypropylmethylcellulose (hypromellose), hydroxymethylcelluloseacetate stearate, polysorbate-80, hydroxyethylcellulose, sodiumalginate, gums, such as, e.g., gum tragacanth and gum acacia, guar gum,xanthans, including xanthan gum, sugars, cellulosics, such as, e.g.,sodium carboxymethylcellulose, methylcellulose, sodiumcarboxymethylcellulose, hydroxypropylmethylcellulose,hydroxyethylcellulose, polysorbate-80, sodium alginate, polyethoxylatedsorbitan monolaurate, polyethoxylated sorbitan monolaurate, povidone andthe like. In some embodiments, useful aqueous suspensions also containone or more polymers as suspending agents. Useful polymers includewater-soluble polymers such as cellulosic polymers, e.g., hydroxypropylmethylcellulose, and water-insoluble polymers such as cross-linkedcarboxyl-containing polymers.

In one embodiment, the present disclosure provides auris-acceptable gelcompositions comprising a therapeutically effective amount of an auralpressure modulating agent in a hydroxyethyl cellulose gel. Hydroxyethylcellulose (HEC) is obtained as a dry powder which is reconstituted inwater or an aqueous buffer solution to give the desired viscosity(generally about 200 cps to about 30,000 cps, corresponding to about 0.2to about 10% HEC). In one embodiment the concentration of HEC is betweenabout 1% and about 15%, about 1% and about 2%, or about 1.5% to about2%.

In other embodiments, the auris-acceptable formulations, including gelformulations and viscosity-enhanced formulations, further includeexcipients, other medicinal or pharmaceutical agents, carriers,adjuvants, such as preserving, stabilizing, wetting or emulsifyingagents, solution promoters, salts, solubilizers, an antifoaming agent,an antioxidant, a dispersing agent, a wetting agent, a surfactant, andcombinations thereof.

Auris-Acceptable Actinic Radiation Curable Gel

In other embodiments, the gel is an actinic radiation curable gel, suchthat following administration to or near the targeted auris structure,use of actinic radiation (or light, including UV light, visible light,or infrared light) the desired gel properties are formed. By way ofexample only, fiber optics are used to provide the actinic radiation soas to form the desired gel properties. In some embodiments, the fiberoptics and the gel administration device form a single unit. In otherembodiments, the fiber optics and the gel administration device areprovided separately.

Auris-Acceptable Solvent Release Gel

In some embodiments, the gel is a solvent release gel such that thedesired gel properties are formed after administration to or near thetargeted auris structure, that is, as the solvent in the injected gelformulation diffuses out the gel, a gel having the desired gelproperties is formed. For example, a formulation that comprises sucroseacetate isobutyrate, a pharmaceutically acceptable solvent, one or moreadditives, and the aural pressure modulating agent is administered at ornear the round window membrane: diffusion of the solvent out of theinjected formulation provides a depot having the desired gel properties.For example, use of a water soluble solvent provides a high viscositydepot when the solvent diffuses rapidly out of the injected formulation.On the other hand, use of a hydrophobic solvent (e.g., benzyl benzoate)provides a less viscous depot. One example of an auris-acceptablesolvent release gel formulation is the SABER™ Delivery System marketedby DURECT Corporation.

Auris-Acceptable In Situ Forming Spongy Material

Also contemplated within the scope of the embodiments is the use of aspongy material, formed in situ in the auris interna or auris media. Insome embodiments, the spongy material is formed from hyaluronic acid orits derivatives. The spongy material is impregnated with a desired auralpressure modulating agent and placed within the auris media so as toprovide controlled release of the aural pressure modulating agent withinthe auris media, or in contact with the round window membrane so as toprovide controlled release of the aural pressure modulating agent intothe auris interna. In some embodiments, the spongy material isbiodegradable.

Round Window Membrane Mucoadhesives

Also contemplated within the scope of the embodiments is the addition ofa round window membrane mucoadhesive with the aural pressure modulatingagent formulations and compositions and devices disclosed herein. Theterm ‘mucoadhesion’ is used for materials that bind to the mucin layerof a biological membrane, such as the external membrane of the 3-layeredround window membrane. To serve as round window membrane mucoadhesivepolymers, the polymers possess some general physiochemical features suchas predominantly anionic hydrophilicity with numerous hydrogen bondforming groups, suitable surface property for wetting mucus/mucosaltissue surfaces or sufficient flexibility to penetrate the mucusnetwork.

Round window membrane mucoadhesive agents that are used with theauris-acceptable formulations include, but are not limited to, at leastone soluble polyvinylpyrrolidone polymer (PVP); a water-swellable, butwater-insoluble, fibrous, cross-linked carboxy-functional polymer; acrosslinked poly(acrylic acid) (e.g. Carbopol® 947P); a carbomerhomopolymer; a carbomer copolymer; a hydrophilic polysaccharide gum,maltodextrin, a cross-linked alignate gum gel, a water-dispersiblepolycarboxylated vinyl polymer, at least two particulate componentsselected from the group consisting of titanium dioxide, silicon dioxide,and clay, or a mixture thereof. The round window membrane mucoadhesiveagent is optionally used in combination with an auris-acceptableviscosity increasing excipient, or used alone to increase theinteraction of the composition with the mucosal layer target oticcomponent. In one non-limiting example, the mucoadhesive agent ismaltodextrin. In some embodiments, the mucoadhesive agent is an alginategum. When used, the round window membrane mucoadhesive characterimparted to the composition is at a level that is sufficient to deliveran effective amount of the aural pressure modulating agent compositionto, for example, the mucosal layer of round window membrane or thecrista fenestrae cochleae in an amount that coats the mucosal membrane,and thereafter deliver the composition to the affected areas, includingby way of example only, the vestibular and/or cochlear structures of theauris interna. When used, the mucoadhesive characteristics of thecompositions provided herein are determined, and using this information(along with the other teachings provided herein), the appropriateamounts are determined. One method for determining sufficientmucoadhesiveness includes monitoring changes in the interaction of thecomposition with a mucosal layer, including but not limited to measuringchanges in residence or retention time of the composition in the absenceand presence of the mucoadhesive excipient.

Mucoadhesive agents have been described, for example, in U.S. Pat. Nos.6,638,521, 6,562,363, 6,509,028, 6,348,502, 6,319,513, 6,306,789,5,814,330, and 4,900,552, each of which is hereby incorporated byreference for such disclosure.

In another non-limiting example, a mucoadhesive agent is, for example,at least two particulate components selected from titanium dioxide,silicon dioxide, and clay, wherein the composition is not furtherdiluted with any liquid prior to administration and the level of silicondioxide, if present, is from about 3% to about 15%, by weight of thecomposition. Silicon dioxide, if present, includes fumed silicondioxide, precipitated silicon dioxide, coacervated silicon dioxide, gelsilicon dioxide, and mixtures thereof. Clay, if present, includes kaolinminerals, serpentine minerals, smectites, illite or a mixture thereof.For example, clay includes laponite, bentonite, hectorite, saponite,montmorillonites or a mixture thereof.

In one non-limiting example, the round window membrane mucoadhesiveagent is maltodextrin. Maltodextrin is a carbohydrate produced by thehydrolysis of starch that is optionally derived from corn, potato, wheator other plant products. Maltodextrin is optionally used either alone orin combination with other round window membrane mucoadhesive agents toimpart mucoadhesive characteristics on the compositions disclosedherein. In one embodiment, a combination of maltodextrin and a carbopolpolymer are used to increase the round window membrane mucoadhesivecharacteristics of the compositions or devices disclosed herein.

In another embodiment, the round window membrane mucoadhesive agent isan alkyl-glycoside and/or a saccharide alkyl ester. As used herein, an“alkyl-glycoside” means a compound comprising any hydrophilic saccharide(e.g. sucrose, maltose, or glucose) linked to a hydrophobic alkyl. Insome embodiments, the round window membrane mucoadhesive agent is analkyl-glycoside wherein the alkyl-glycoside comprises a sugar linked toa hydrophobic alkyl (e.g., an alkyl comprising about 6 to about 25carbon atoms) by an amide linkage, an amine linkage, a carbamatelinkage, an ether linkage, a thioether linkage, an ester linkage, athioester linkage, a glycosidic linkage, a thioglycosidic linkage,and/or a ureide linkage. In some embodiments, the round window membranemucoadhesive agent is a hexyl-, heptyl-, octyl-, nonyl-, decyl-,undecyl-, dodecyl-, tridecyl-, tetradecyl, pentadecyl-, hexadecyl-,heptadecyl-, and octadecyl α- or β-D-maltoside; hexyl-, heptyl-, octyl-,nonyl-, decyl-, undecyl-, dodecyl-, tridecyl-, tetradecyl, pentadecyl-,hexadecyl-, heptadecyl-, and octadecyl α- or β-D-glucoside; hexyl-,heptyl-, octyl-, nonyl-, decyl-, undecyl-, dodecyl-, tridecyl-,tetradecyl, pentadecyl-, hexadecyl-, heptadecyl-, and octadecyl α- orβ-D-sucroside; hexyl-, heptyl-, octyl-, dodecyl-, tridecyl-, andtetradecyl-β-D-thiomaltoside; heptyl- or octyl-1-thio-α- orβ-D-glucopyranoside; alkyl thiosucroses; alkyl maltotriosides; longchain aliphatic carbonic acid amides of sucrose β-amino-alkyl ethers;derivatives of palatinose or isomaltamine linked by an amide linkage toan alkyl chain and derivatives of isomaltamine linked by urea to analkyl chain; long chain aliphatic carbonic acid ureides of sucroseβ-amino-alkyl ethers and long chain aliphatic carbonic acid amides ofsucrose β-amino-alkyl ethers. In some embodiments, the round windowmembrane mucoadhesive agent is an alkyl-glycoside wherein the alkylglycoside is maltose, sucrose, glucose, or a combination thereof linkedby a glycosidic linkage to an alkyl chain of 9-16 carbon atoms (e.g.,nonyl-, decyl-, dodecyl- and tetradecyl sucroside; nonyl-, decyl-,dodecyl- and tetradecyl glucoside; and nonyl-, decyl-, dodecyl- andtetradecyl maltoside). In some embodiments, the round window membranemucoadhesive agent is an alkyl-glycoside wherein the alkyl glycoside isdodecylmaltoside, tridecylmalto side, and tetradecylmaltoside.

In some embodiments, the round window membrane mucoadhesive agent is analkyl-glycoside wherein the alkyl-glycoside is a disaccharide with atleast one glucose. In some embodiments, the auris acceptable penetrationenhancer is a surfactant comprisingα-D-glucopyranosyl-β-glycopyranoside,n-Dodecyl-4-O-α-D-glucopyranosyl-β-glycopyranoside, and/orn-tetradecyl-4-O-α-D-glucopyranosyl-β-glycopyranoside. In someembodiments, the round window membrane mucoadhesive agent is analkyl-glycoside wherein the alkyl-glycoside has a critical miscelleconcentration (CMC) of less than about 1 mM in pure water or in aqueoussolutions. In some embodiments, the round window membrane mucoadhesiveagent is an alkyl-glycoside wherein an oxygen atom within thealkyl-glycoside is substituted with a sulfur atom. In some embodiments,the round window membrane mucoadhesive agent is an alkyl-glycosidewherein the alkylglycoside is the β anomer. In some embodiments, theround window membrane mucoadhesive agent is an alkyl-glycoside whereinthe alkylglycoside comprises 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%,98%, 99%, 99.1%, 99.5%, or 99.9% of the β anomer.

Auris-Acceptable Controlled Release Particles

Aural pressure modulating agents and/or other pharmaceutical agentsdisclosed herein are optionally incorporated within controlled releaseparticles, lipid complexes, liposomes, nanoparticles, microparticles,microspheres, coacervates, nanocapsules or other agents which enhance orfacilitate the localized delivery of the aural pressure modulatingagent. In some embodiments, a single enhanced viscosity formulation isused, in which at least one aural pressure modulating agent is present,while in other embodiments, a pharmaceutical formulation that comprisesa mixture of two or more distinct enhanced viscosity formulations isused, in which at least one aural pressure modulating agent is present.In some embodiments, combinations of sols, gels and/or biocompatiblematrices is also employed to provide desirable characteristics of thecontrolled release aural pressure modulating compositions orformulations. In certain embodiments, the controlled release auralpressure modulating formulations or compositions are cross-linked by oneor more agents to alter or improve the properties of the composition.

Examples of microspheres relevant to the pharmaceutical formulationsdisclosed herein include: Luzzi, L. A., J. Pharm. Psy. 59:1367 (1970);U.S. Pat. No. 4,530,840; Lewis, D. H., “Controlled Release of BioactiveAgents from Lactides/Glycolide Polymers” in Biodegradable Polymers asDrug Delivery Systems, Chasin, M. and Langer, R., eds., Marcel Decker(1990); U.S. Pat. No. 4,675,189; Beck et al., “Poly(lactic acid) andPoly(lactic acid-co-glycolic acid) Contraceptive Delivery Systems,” inLong Acting Steroid Contraception, Mishell, D. R., ed., Raven Press(1983); U.S. Pat. No. 4,758,435; U.S. Pat. No. 3,773,919; U.S. Pat. No.4,474,572. Examples of protein therapeutics formulated as microspheresinclude: U.S. Pat. No. 6,458,387; U.S. Pat. No. 6,268,053; U.S. Pat. No.6,090,925; U.S. Pat. No. 5,981,719; and U.S. Pat. No. 5,578,709, and areherein incorporated by reference for such disclosure.

Microspheres usually have a spherical shape, although irregularly-shapedmicroparticles are possible. Microspheres may vary in size, ranging fromsubmicron to 1000 micron diameters. Microspheres suitable for use withthe auris-acceptable formulations disclosed herein are submicron to 250micron diameter microspheres, allowing administration by injection witha standard gauge needle. The auris-acceptable microspheres are preparedby any method which produces microspheres in a size range acceptable foruse in an injectable composition. Injection is optionally accomplishedwith standard gauge needles used for administering liquid compositions.

Suitable examples of polymeric matrix materials for use in theauris-acceptable controlled release particles herein includepoly(glycolic acid), poly-d,l-lactic acid, poly-l-lactic acid,copolymers of the foregoing, poly(aliphatic carboxylic acids),copolyoxalates, polycaprolactone, polydioxonene, poly(orthocarbonates),poly(acetals), poly(lactic acid-caprolactone), polyorthoesters,poly(glycolic acid-caprolactone), polydioxonene, polyanhydrides,polyphosphazines, and natural polymers including albumin, casein, andsome waxes, such as, glycerol mono- and distearate, and the like.Various commercially available poly(lactide-co-glycolide) materials(PLGA) are optionally used in the method disclosed herein. For example,poly(d,l-lactic-co-glycolic acid) is commercially available fromBoehringer-Ingelheim as RESOMER RG 503 H. This product has a molepercent composition of 50% lactide and 50% glycolide. These copolymersare available in a wide range of molecular weights and ratios of lacticacid to glycolic acid. One embodiment includes the use of the polymerpoly(d,l-lactide-co-glycolide). The molar ratio of lactide to glycolidein such a copolymer includes the range of from about 95:5 to about50:50.

The molecular weight of the polymeric matrix material is of someimportance. The molecular weight should be high enough so that it formssatisfactory polymer coatings, i.e., the polymer should be a good filmformer. Usually, a satisfactory molecular weight is in the range of5,000 to 500,000 daltons. The molecular weight of a polymer is alsoimportant from the point of view that molecular weight influences thebiodegradation rate of the polymer. For a diffusional mechanism of drugrelease, the polymer should remain intact until all of the drug isreleased from the microparticles and then degrade. The drug is alsoreleased from the microparticles as the polymeric excipient bioerodes.By an appropriate selection of polymeric materials a microsphereformulation is made such that the resulting microspheres exhibit bothdiffusional release and biodegradation release properties. This isuseful in affording multiphasic release patterns.

A variety of methods are known by which compounds are encapsulated inmicrospheres. In these methods, the aural pressure modulating agent isgenerally dispersed or emulsified, using stirrers, agitators, or otherdynamic mixing techniques, in a solvent containing a wall-formingmaterial. Solvent is then removed from the microspheres, and thereafterthe microsphere product is obtained.

In one embodiment, controlled release aural pressure modulatingformulations are made through the incorporation of the aural pressuremodulating agents and/or other pharmaceutical agents into ethylene-vinylacetate copolymer matrices. (See U.S. Pat. No. 6,083,534, incorporatedherein for such disclosure). In another embodiment, aural pressuremodulating agents are incorporated into poly(lactic-glycolic acid) orpoly-L-lactic acid microspheres. Id. In yet another embodiment, theaural pressure modulating agents are encapsulated into alginatemicrospheres. (See U.S. Pat. No. 6,036,978, incorporated herein for suchdisclosure). Biocompatible methacrylate-based polymers to encapsulatethe aural pressure modulating compounds or compositions are optionallyused in the formulations and methods disclosed herein. A wide range ofmethacrylate-based polymer systems are commercially available, such asthe EUDRAGIT polymers marketed by Evonik. One useful aspect ofmethacrylate polymers is that the properties of the formulation arevaried by incorporating various co-polymers. For example, poly(acrylicacid-co-methylmethacrylate) microparticles exhibit enhanced mucoadhesionproperties as the carboxylic acid groups in the poly(acrylic acid) formhydrogen bonds with mucin (Park et al, Pharm. Res. (1987) 4(6):457-464). Variation of the ratio between acrylic acid andmethylmethacrylate monomers serves to modulate the properties of theco-polymer. Methacrylate-based microparticles have also been used inprotein therapeutic formulations (Naha et al, Journal ofMicroencapsulation 4 Feb., 2008 (online publication)). In oneembodiment, the enhanced viscosity auris-acceptable formulationsdescribed herein comprises aural pressure modulating microsphereswherein the microspheres are formed from a methacrylate polymer orcopolymer. In an additional embodiment, the enhanced viscosityformulation described herein comprises aural pressure modulatingmicrospheres wherein the microspheres are mucoadhesive. Other controlledrelease systems, including incorporation or deposit of polymericmaterials or matrices onto solid or hollow spheres containing auralpressure modulating agents, are also explicitly contemplated within theembodiments disclosed herein. The types of controlled release systemsavailable without significantly losing activity of the aural pressuremodulating agent are determined using the teachings, examples, andprinciples disclosed herein

An example of a conventional microencapsulation process forpharmaceutical preparations is shown in U.S. Pat. No. 3,737,337,incorporated herein by reference for such disclosure. The aural pressuremodulating substances to be encapsulated or embedded are dissolved ordispersed in the organic solution of the polymer (phase A), usingconventional mixers, including (in the preparation of dispersion)vibrators and high-speed stirrers, etc. The dispersion of phase (A),containing the core material in solution or in suspension, is carriedout in the aqueous phase (B), again using conventional mixers, such ashigh-speed mixers, vibration mixers, or even spray nozzles, in whichcase the particle size of the microspheres will be determined not onlyby the concentration of phase (A), but also by the emulsate ormicrosphere size. With conventional techniques for themicroencapsulation of aural pressure modulating agents, the microspheresform when the solvent containing an active agent and a polymer isemulsified or dispersed in an immiscible solution by stirring,agitating, vibrating, or some other dynamic mixing technique, often fora relatively long period of time.

Methods for the construction of microspheres are also described in U.S.Pat. No. 4,389,330, and U.S. Pat. No. 4,530,840, incorporated herein byreference for such disclosure. The desired aural pressure modulatingagent is dissolved or dispersed in an appropriate solvent. To theagent-containing medium is added the polymeric matrix material in anamount relative to the active ingredient which gives a product of thedesired loading of active agent. Optionally, all of the ingredients ofthe aural pressure modulating microsphere product can be blended in thesolvent medium together. Suitable solvents for the agent and thepolymeric matrix material include organic solvents such as acetone,halogenated hydrocarbons such as chloroform, methylene chloride and thelike, aromatic hydrocarbon compounds, halogenated aromatic hydrocarboncompounds, cyclic ethers, alcohols, ethyl acetate and the like.

The mixture of ingredients in the solvent is emulsified in acontinuous-phase processing medium; the continuous-phase medium beingsuch that a dispersion of microdroplets containing the indicatedingredients is formed in the continuous-phase medium. Naturally, thecontinuous-phase processing medium and the organic solvent must beimmiscible, and includes water although nonaqueous media such as xyleneand toluene and synthetic oils and natural oils are optionally used.Optionally, a surfactant is added to the continuous-phase processingmedium to prevent the microparticles from agglomerating and to controlthe size of the solvent microdroplets in the emulsion. A preferredsurfactant-dispersing medium combination is a 1 to 10 wt. % poly (vinylalcohol) in water mixture. The dispersion is formed by mechanicalagitation of the mixed materials. An emulsion is optionally formed byadding small drops of the active agent-wall forming material solution tothe continuous phase processing medium. The temperature during theformation of the emulsion is not especially critical but influences thesize and quality of the microspheres and the solubility of the drug inthe continuous phase. It is desirable to have as little of the agent inthe continuous phase as possible. Moreover, depending on the solvent andcontinuous-phase processing medium employed, the temperature must not betoo low or the solvent and processing medium will solidify or theprocessing medium will become too viscous for practical purposes, or toohigh that the processing medium will evaporate, or that the liquidprocessing medium will not be maintained. Moreover, the temperature ofthe medium cannot be so high that the stability of the particular agentbeing incorporated in the microspheres is adversely affected.Accordingly, the dispersion process is conducted at any temperaturewhich maintains stable operating conditions, which preferred temperaturebeing about 15° C. to 60° C., depending upon the drug and excipientselected.

The dispersion which is formed is a stable emulsion and from thisdispersion the organic solvent immiscible fluid is optionally partiallyremoved in the first step of the solvent removal process. The solvent isremoved by techniques such as heating, the application of a reducedpressure or a combination of both. The temperature employed to evaporatesolvent from the microdroplets is not critical, but should not be thathigh that it degrades the aural pressure modulating agent employed inthe preparation of a given microparticle, nor should it be so high as toevaporate solvent at such a rapid rate to cause defects in the wallforming material. Generally, from 5 to 75%, of the solvent is removed inthe first solvent removal step.

After the first stage, the dispersed microparticles in the solventimmiscible fluid medium are isolated from the fluid medium by anyconvenient means of separation. Thus, for example, the fluid is decantedfrom the microsphere or the microsphere suspension is filtered. Stillother, various combinations of separation techniques are used ifdesired.

Following the isolation of the microspheres from the continuous-phaseprocessing medium, the remainder of the solvent in the microspheres isremoved by extraction. In this step, the microspheres are suspended inthe same continuous-phase processing medium used in step one, with orwithout surfactant, or in another liquid. The extraction medium removesthe solvent from the microspheres and yet does not dissolve themicrospheres. During the extraction, the extraction medium withdissolved solvent is optionally removed and replaced with freshextraction medium. This is best done on a continual basis. The rate ofextraction medium replenishment of a given process is a variable whichis determined at the time the process is performed and, therefore, noprecise limits for the rate must be predetermined. After the majority ofthe solvent has been removed from the microspheres, the microspheres aredried by exposure to air or by other conventional drying techniques suchas vacuum drying, drying over a desiccant, or the like. This process isvery efficient in encapsulating the aural pressure modulating agentsince core loadings of up to 80 wt. %, preferably up to 60 wt. % areobtained.

Alternatively, controlled release microspheres containing an auralpressure modulating agent is prepared through the use of static mixers.Static or motionless mixers consist of a conduit or tube in which isreceived a number of static mixing agents. Static mixers providehomogeneous mixing in a relatively short length of conduit, and in arelatively short period of time. With static mixers, the fluid movesthrough the mixer, rather than some part of the mixer, such as a blade,moving through the fluid.

A static mixer is optionally used to create an emulsion. When using astatic mixer to form an emulsion, several factors determine emulsionparticle size, including the density and viscosity of the varioussolutions or phases to be mixed, volume ratio of the phases, interfacialtension between the phases, static mixer parameters (conduit diameter;length of mixing element; number of mixing elements), and linearvelocity through the static mixer. Temperature is a variable because itaffects density, viscosity, and interfacial tension. The controllingvariables are linear velocity, sheer rate, and pressure drop per unitlength of static mixer.

In order to create microspheres containing an aural pressure modulatingagent using a static mixer process, an organic phase and an aqueousphase are combined. The organic and aqueous phases are largely orsubstantially immiscible, with the aqueous phase constituting thecontinuous phase of the emulsion. The organic phase includes an auralpressure modulating agent as well as a wall-forming polymer or polymericmatrix material. The organic phase is prepared by dissolving an auralpressure modulating agent in an organic or other suitable solvent, or byforming a dispersion or an emulsion containing the aural pressuremodulating agent. The organic phase and the aqueous phase are pumped sothat the two phases flow simultaneously through a static mixer, therebyforming an emulsion which comprises microspheres containing the auralpressure modulating agent encapsulated in the polymeric matrix material.The organic and aqueous phases are pumped through the static mixer intoa large volume of quench liquid to extract or remove the organicsolvent. Organic solvent is optionally removed from the microsphereswhile they are washing or being stirred in the quench liquid. After themicrospheres are washed in a quench liquid, they are isolated, asthrough a sieve, and dried.

In one embodiment, microspheres are prepared using a static mixer. Theprocess is not limited to the solvent extraction technique discussedabove, but is used with other encapsulation techniques. For example, theprocess is optionally used with a phase separation encapsulationtechnique. To do so, an organic phase is prepared that comprises anaural pressure modulating agent suspended or dispersed in a polymersolution. The non-solvent second phase is free from solvents for thepolymer and active agent. A preferred non-solvent second phase issilicone oil. The organic phase and the non-solvent phase are pumpedthrough a static mixer into a non-solvent quench liquid, such asheptane. The semi-solid particles are quenched for complete hardeningand washing. The process of microencapsulation includes spray drying,solvent evaporation, a combination of evaporation and extraction, andmelt extrusion.

In another embodiment, the microencapsulation process involves the useof a static mixer with a single solvent. This process is described indetail in U.S. application Ser. No. 08/338,805, herein incorporated byreference for such disclosure. An alternative process involves the useof a static mixer with co-solvents. In this process, biodegradablemicrospheres comprising a biodegradable polymeric binder and an auralpressure modulating agent are prepared, which comprises a blend of atleast two substantially non-toxic solvents, free of halogenatedhydrocarbons to dissolve both the agent and the polymer. The solventblend containing the dissolved agent and polymer is dispersed in anaqueous solution to form droplets. The resulting emulsion is then addedto an aqueous extraction medium preferably containing at least one ofthe solvents of the blend, whereby the rate of extraction of eachsolvent is controlled, whereupon the biodegradable microspherescontaining the pharmaceutically active agent are formed. This processhas the advantage that less extraction medium is required because thesolubility of one solvent in water is substantially independent of theother and solvent selection is increased, especially with solvents thatare particularly difficult to extract.

Nanoparticles are also contemplated for use with the aural pressuremodulating agents disclosed herein. Nanoparticles are materialstructures of about 100 nm or less in size. One use of nanoparticles inpharmaceutical formulations is the formation of suspensions as theinteraction of the particle surface with solvent is strong enough toovercome differences in density. Nanoparticle suspensions are sterilizedas the nanoparticles are small enough to be subjected to sterilizingfiltration (see, e.g., U.S. Pat. No. 6,139,870, herein incorporated byreference for such disclosure). Nanoparticles comprise at least onehydrophobic, water-insoluble and water-indispersible polymer orcopolymer emulsified in a solution or aqueous dispersion of surfactants,phospholipids or fatty acids. The aural pressure modulating agent isoptionally introduced with the polymer or the copolymer into thenanoparticles.

Lipid nanocapsules as controlled release structures, as well forpenetrating the round window membrane and reaching auris interna and/orauris media targets, is also contemplated herein. Lipid nanocapsules areoptionally formed by emulsifying capric and caprylic acid triglycerides(Labrafac WL 1349; avg. mw 512), soybean lecithin (LIPOID® S75-3; 69%phosphatidylcholine and other phospholipids), surfactant (for example,Solutol HS15), a mixture of polyethylene glycol 660 hydroxystearate andfree polyethylene glycol 660; NaCl and water. The mixture is stirred atroom temperature to obtain an oil emulsion in water. After progressiveheating at a rate of 4° C./min under magnetic stirring, a short intervalof transparency should occur close to 70° C., and the inverted phase(water droplets in oil) obtained at 85° C. Three cycles of cooling andheating is then applied between 85° C. and 60° C. at the rate of 4°C./min, and a fast dilution in cold water at a temperature close to 0°C. to produce a suspension of nanocapsules. To encapsulate the auralpressure modulating agents, the agent is optionally added just prior tothe dilution with cold water.

Aural pressure modulating agents are also inserted into the lipidnanocapsules by incubation for 90 minutes with an aqueous micellarsolution of the auris active agent. The suspension is then vortexedevery 15 minutes, and then quenched in an ice bath for 1 minute.

Suitable auris-acceptable surfactants are, by way of example, cholicacid or taurocholic acid salts. Taurocholic acid, the conjugate formedfrom cholic acid and taurine, is a fully metabolizable sulfonic acidsurfactant. An analog of taurocholic acid, tauroursodeoxycholic acid(TUDCA), is a naturally occurring bile acid and is a conjugate oftaurine and ursodeoxycholic acid (UDCA). Other naturally occurringanionic (e.g., galactocerebroside sulfate), neutral (e.g.,lactosylceramide) or zwitterionic surfactants (e.g., sphingomyelin,phosphatidyl choline, palmitoyl carnitine) are optionally used toprepare nanoparticles.

The auris-acceptable phospholipids are chosen, by way of example, fromnatural, synthetic or semi-synthetic phospholipids; lecithins(phosphatidylcholine) such as, for example, purified egg or soyalecithins (lecithin E100, lecithin E80 and phospholipons, for examplephospholipon 90), phosphatidylethanolamine, phosphatidylserine,phosphatidylinositol, phosphatidylglycerol,dipalmitoylphosphatidylcholine, dipalmitoylglycerophosphatidylcholine,dimyristoylphosphatidylcholine, distearoylphosphatidylcholine andphosphatidic acid or mixtures thereof are used more particularly.

Fatty acids for use with the auris-acceptable formulations are chosenfrom, by way of example, lauric acid, mysristic acid, palmitic acid,stearic acid, isostearic acid, arachidic acid, behenic acid, oleic acid,myristoleic acid, palmitoleic acid, linoleic acid, alpha-linoleic acid,arachidonic acid, eicosapentaenoic acid, erucic acid, docosahexaenoicacid, and the like.

Suitable auris-acceptable surfactants are selected from known organicand inorganic pharmaceutical excipients. Such excipients include variouspolymers, low molecular weight oligomers, natural products, andsurfactants. Preferred surface modifiers include nonionic and ionicsurfactants. Two or more surface modifiers are used in combination.

Representative examples of auris-acceptable surfactants include cetylpyridinium chloride, gelatin, casein, lecithin (phosphatides), dextran,glycerol, gum acacia, cholesterol, tragacanth, stearic acid, calciumstearate, glycerol monostearate, cetostearyl alcohol, cetomacrogolemulsifying wax, sorbitan esters, polyoxyethylene alkyl ethers,polyoxyethylene castor oil derivatives, polyoxyethylene sorbitan fattyacid esters; dodecyl trimethyl ammonium bromide,polyoxyethylenestearates, colloidal silicon dioxide, phosphates, sodiumdodecylsulfate, carboxymethylcellulose calcium, hydroxypropyl cellulose(HPC, HPC-SL, and HPC-L), hydroxypropyl methylcellulose (HPMC),carboxymethylcellulose sodium, methylcellulose, hydroxyethylcellulose,hydroxypropylcellulose, hydroxypropylmethyl-cellulose phthalate,noncrystalline cellulose, magnesium aluminum silicate, triethanolamine,polyvinyl alcohol (PVA), polyvinylpyrrolidone (PVP),4-(1,1,3,3-tetaamethylbutyl)-phenol polymer with ethylene oxide andformaldehyde (also known as tyloxapol, superione, and triton),poloxamers, poloxamines, a charged phospholipid such as dimyristoylphophatidyl glycerol, dioctylsulfosuccinate (DOSS); Tetronic® 1508,dialkylesters of sodium sulfosuccinic acid, Duponol P, Tritons X-200,Crodestas F-110, p-isononylphenoxypoly-(glycidol), Crodestas SL-40(Croda, Inc.); and SA9OHCO, which isC₁₈H₃₇CH₂(CON(CH₃)—CH₂(CHOH)₄(CH₂OH)₂ (Eastman Kodak Co.);decanoyl-N-methylglucamide; n-decyl β-D-glucopyranoside; n-decylβ-D-maltopyranoside; n-dodecyl β-D-glucopyranoside; n-dodecylβ-D-maltoside; heptanoyl-N-methylglucamide;n-heptyl-β-D-glucopyranoside; n-heptyl β-D-thioglucoside; n-hexylβ-D-glucopyranoside; nonanoyl-N-methylglucamide; n-noylβ-D-glucopyranoside; octanoyl-N-methylglucamide;n-octyl-β-D-glucopyranoside; octyl β-D-thioglucopyranoside; and thelike. Most of these surfactants are known pharmaceutical excipients andare described in detail in the Handbook of Pharmaceutical Excipients,published jointly by the American Pharmaceutical Association and ThePharmaceutical Society of Great Britain (The Pharmaceutical Press,1986), specifically incorporated by reference for such disclosure.

The hydrophobic, water-insoluble and water-indispersible polymer orcopolymer may be chosen from biocompatible and biodegradable polymers,for example lactic or glycolic acid polymers and copolymers thereof, orpolylactic/polyethylene (or polypropylene) oxide copolymers, preferablywith molecular weights of between 1000 and 200,000, polyhydroxybutyricacid polymers, polylactones of fatty acids containing at least 12 carbonatoms, or polyanhydrides.

The nanoparticles may be obtained by coacervation, or by the techniqueof evaporation of solvent, from an aqueous dispersion or solution ofphospholipids and of an oleic acid salt into which is added animmiscible organic phase comprising the active principle and thehydrophobic, water-insoluble and water-indispersible polymer orcopolymer. The mixture is pre-emulsified and then subjected tohomogenization and evaporation of the organic solvent to obtain anaqueous suspension of very small-sized nanoparticles.

A variety of methods are optionally employed to fabricate the auralpressure modulating nanoparticles that are within the scope of theembodiments. These methods include vaporization methods, such as freejet expansion, laser vaporization, spark erosion, electro explosion andchemical vapor deposition; physical methods involving mechanicalattrition (e.g., “pearlmilling” technology, Elan Nanosystems), supercritical CO2 and interfacial deposition following solvent displacement.In one embodiment, the solvent displacement method is used. The size ofnanoparticles produced by this method is sensitive to the concentrationof polymer in the organic solvent; the rate of mixing; and to thesurfactant employed in the process. Continuous flow mixers provide thenecessary turbulence to ensure small particle size. One type ofcontinuous flow mixing device that is optionally used to preparenanoparticles has been described (Hansen et al J Phys Chem 92, 2189-96,1988). In other embodiments, ultrasonic devices, flow throughhomogenizers or supercritical CO2 devices may be used to preparenanoparticles.

If suitable nanoparticle homogeneity is not obtained on directsynthesis, then size-exclusion chromatography is used to produce highlyuniform drug-containing particles that are freed of other componentsinvolved in their fabrication. Size-exclusion chromatography (SEC)techniques, such as gel-filtration chromatography, is used to separateparticle-bound aural pressure modulating agent or other pharmaceuticalcompound from free aural pressure modulating agent or otherpharmaceutical compound, or to select a suitable size range of auralpressure modulator-containing nanoparticles. Various SEC media, such asSuperdex 200, Superose 6, Sephacryl 1000 are commercially available andare employed for the size-based fractionation of such mixtures.Additionally, nanoparticles are optionally purified by centrifugation,membrane filtration and by use of other molecular sieving devices,crosslinked gels/materials and membranes.

Auris-Acceptable Cyclodextrin and Other Stabilizing Formulations

In a specific embodiment, the auris-acceptable formulationsalternatively comprises a cyclodextrin. Cyclodextrins are cyclicoligosaccharides containing 6, 7, or 8 glucopyranose units, referred toas α-cyclodextrin, β-cyclodextrin, or γ-cyclodextrin respectively.Cyclodextrins have a hydrophilic exterior, which enhances water-soluble,and a hydrophobic interior which forms a cavity. In an aqueousenvironment, hydrophobic portions of other molecules often enter thehydrophobic cavity of cyclodextrin to form inclusion compounds.Additionally, cyclodextrins are also capable of other types ofnonbonding interactions with molecules that are not inside thehydrophobic cavity. Cyclodextrins have three free hydroxyl groups foreach glucopyranose unit, or 18 hydroxyl groups on α-cyclodextrin, 21hydroxyl groups on β-cyclodextrin, and 24 hydroxyl groups onγ-cyclodextrin. One or more of these hydroxyl groups can be reacted withany of a number of reagents to form a large variety of cyclodextrinderivatives, including hydroxypropyl ethers, sulfonates, andsulfoalkylethers. Shown below is the structure of β-cyclodextrin and thehydroxypropyl-β-cyclodextrin (HPβCD).

In some embodiments, the use of cyclodextrins in the pharmaceuticalcompositions described herein improves the solubility of the drug.Inclusion compounds are involved in many cases of enhanced solubility;however other interactions between cyclodextrins and insoluble compoundsalso improves solubility. Hydroxypropyl-β-cyclodextrin (HPβCD) iscommercially available as a pyrogen free product. It is a nonhygroscopicwhite powder that readily dissolves in water. HPβCD is thermally stableand does not degrade at neutral pH. Thus, cyclodextrins improve thesolubility of a therapeutic agent in a composition or formulation.Accordingly, in some embodiments, cyclodextrins are included to increasethe solubility of the auris-acceptable aural pressure modulating agentswithin the formulations described herein. In other embodiments,cyclodextrins in addition serve as controlled release excipents withinthe formulations described herein.

By way of example only, cyclodextrin derivatives for use includeα-cyclodextrin, β-cyclodextrin, γ-cyclodextrin, hydroxyethylβ-cyclodextrin, hydroxypropyl γ-cyclodextrin, sulfated β-cyclodextrin,sulfated α-cyclodextrin, sulfobutyl ether β-cyclodextrin.

The concentration of the cyclodextrin used in the compositions andmethods disclosed herein varies according to the physiochemicalproperties, pharmacokinetic properties, side effect or adverse events,formulation considerations, or other factors associated with thetherapeutically active agent, or a salt or prodrug thereof, or with theproperties of other excipients in the composition. Thus, in certaincircumstances, the concentration or amount of cyclodextrin used inaccordance with the compositions and methods disclosed herein will vary,depending on the need. When used, the amount of cyclodextrins needed toincrease solubility of the aural pressure modulating agent and/orfunction as a controlled release excipient in any of the formulationsdescribed herein is selected using the principles, examples, andteachings described herein.

Other stabilizers that are useful in the auris-acceptable formulationsdisclosed herein include, for example, fatty acids, fatty alcohols,alcohols, long chain fatty acid esters, long chain ethers, hydrophilicderivatives of fatty acids, polyvinyl pyrrolidones, polyvinyl ethers,polyvinyl alcohols, hydrocarbons, hydrophobic polymers,moisture-absorbing polymers, and combinations thereof. In someembodiments, amide analogues of stabilizers are also used. In furtherembodiments, the chosen stabilizer changes the hydrophobicity of theformulation (e.g., oleic acid, waxes), or improves the mixing of variouscomponents in the formulation (e.g., ethanol), controls the moisturelevel in the formula (e.g., PVP or polyvinyl pyrrolidone), controls themobility of the phase (substances with melting points higher than roomtemperature such as long chain fatty acids, alcohols, esters, ethers,amides etc. or mixtures thereof; waxes), and/or improves thecompatibility of the formula with encapsulating materials (e.g., oleicacid or wax). In another embodiment some of these stabilizers are usedas solvents/co-solvents (e.g., ethanol). In other embodiments,stabilizers are present in sufficient amounts to inhibit the degradationof the aural pressure modulating agent. Examples of such stabilizingagents, include, but are not limited to: (a) about 0.5% to about 2% w/vglycerol, (b) about 0.1% to about 1% w/v methionine, (c) about 0.1% toabout 2% w/v monothioglycerol, (d) about 1 mM to about 10 mM EDTA, (e)about 0.01% to about 2% w/v ascorbic acid, (f) 0.003% to about 0.02% w/vpolysorbate 80, (g) 0.001% to about 0.05% w/v. polysorbate 20, (h)arginine, (i) heparin, (j) dextran sulfate, (k) cyclodextrins, (l)pentosan polysulfate and other heparinoids, (m) divalent cations such asmagnesium and zinc; or (n) combinations thereof.

Additional useful aural pressure modulating agent auris-acceptableformulations include one or more anti-aggregation additives to enhancestability of aural pressure modulating agent formulations by reducingthe rate of protein aggregation. The anti-aggregation additive selecteddepends upon the nature of the conditions to which the aural pressuremodulating agents, for example aural pressure modulating agentantibodies are exposed. For example, certain formulations undergoingagitation and thermal stress require a different anti-aggregationadditive than a formulation undergoing lyophilization andreconstitution. Useful anti-aggregation additives include, by way ofexample only, urea, guanidinium chloride, simple amino acids such asglycine or arginine, sugars, polyalcohols, polysorbates, polymers suchas polyethylene glycol and dextrans, alkyl saccharides, such as alkylglycoside, and surfactants.

Other useful formulations optionally include one or moreauris-acceptable antioxidants to enhance chemical stability whererequired. Suitable antioxidants include, by way of example only,ascorbic acid, methionine, sodium thiosulfate and sodium metabisulfite.In one embodiment, antioxidants are selected from metal chelatingagents, thiol containing compounds and other general stabilizing agents.

Still other useful compositions include one or more auris-acceptablesurfactants to enhance physical stability or for other purposes.Suitable nonionic surfactants include, but are not limited to,polyoxyethylene fatty acid glycerides and vegetable oils, e.g.,polyoxyethylene (60) hydrogenated castor oil; and polyoxyethylenealkylethers and alkylphenyl ethers, e.g., octoxynol 10, octoxynol 40.

In some embodiments, the auris-acceptable pharmaceutical formulationsdescribed herein are stable with respect to compound degradation over aperiod of any of at least about 1 day, at least about 2 days, at leastabout 3 days, at least about 4 days, at least about 5 days, at leastabout 6 days, at least about 1 week, at least about 2 weeks, at leastabout 3 weeks, at least about 4 weeks, at least about 5 weeks, at leastabout 6 weeks, at least about 7 weeks, at least about 8 weeks, at leastabout 3 months, at least about 4 months, at least about 5 months, or atleast about 6 months. In other embodiments, the formulations describedherein are stable with respect to compound degradation over a period ofat least about 1 week. Also described herein are formulations that arestable with respect to compound degradation over a period of at leastabout 1 month.

In other embodiments, an additional surfactant (co-surfactant) and/orbuffering agent is combined with one or more of the pharmaceuticallyacceptable vehicles previously described herein so that the surfactantand/or buffering agent maintains the product at an optimal pH forstability. Suitable co-surfactants include, but are not limited to: a)natural and synthetic lipophilic agents, e.g., phospholipids,cholesterol, and cholesterol fatty acid esters and derivatives thereof;b) nonionic surfactants, which include for example, polyoxyethylenefatty alcohol esters, sorbitan fatty acid esters (Spans),polyoxyethylene sorbitan fatty acid esters (e.g., polyoxyethylene (20)sorbitan monooleate (Tween 80), polyoxyethylene (20) sorbitanmonostearate (Tween 60), polyoxyethylene (20) sorbitan monolaurate(Tween 20) and other Tweens, sorbitan esters, glycerol esters, e.g.,Myrj and glycerol triacetate (triacetin), polyethylene glycols, cetylalcohol, cetostearyl alcohol, stearyl alcohol, polysorbate 80,poloxamers, poloxamines, polyoxyethylene castor oil derivatives (e.g.,Cremophor® RH40, Cremphor A25, Cremphor A20, Cremophor® EL) and otherCremophors, sulfosuccinates, alkyl sulphates (SLS); PEG glyceryl fattyacid esters such as PEG-8 glyceryl caprylate/caprate (Labrasol), PEG-4glyceryl caprylate/caprate (Labrafac Hydro WL 1219), PEG-32 glyceryllaurate (Gelucire 444/14), PEG-6 glyceryl mono oleate (Labrafil M 1944CS), PEG-6 glyceryl linoleate (Labrafil M 2125 CS); propylene glycolmono- and di-fatty acid esters, such as propylene glycol laurate,propylene glycol caprylate/caprate; Brij® 700, ascorbyl-6-palmitate,stearylamine, sodium lauryl sulfate, polyoxethyleneglyceroltriricinoleate, and any combinations or mixtures thereof; c) anionicsurfactants include, but are not limited to, calciumcarboxymethylcellulose, sodium carboxymethylcellulose, sodiumsulfosuccinate, dioctyl, sodium alginate, alkyl polyoxyethylenesulfates, sodium lauryl sulfate, triethanolamine stearate, potassiumlaurate, bile salts, and any combinations or mixtures thereof; and d)cationic surfactants such as cetyltrimethylammonium bromide, andlauryldimethylbenzyl-ammonium chloride.

In a further embodiment, when one or more co-surfactants are utilized inthe auris-acceptable formulations of the present disclosure, they arecombined, e.g., with a pharmaceutically acceptable vehicle and ispresent in the final formulation, e.g., in an amount ranging from about0.1% to about 20%, from about 0.5% to about 10%.

In one embodiment, the surfactant has an HLB value of 0 to 20. Inadditional embodiments, the surfactant has an HLB value of 0 to 3, of 4to 6, of 7 to 9, of 8 to 18, of 13 to 15, of 10 to 18.

In one embodiment, diluents are also used to stabilize the auralpressure modulating agent or other pharmaceutical compounds because theyprovide a more stable environment. Salts dissolved in buffered solutions(which also can provide pH control or maintenance) are utilized asdiluents, including, but not limited to a phosphate buffered salinesolution. In other embodiments, the gel formulation is isotonic with theendolymph or the perilymph: depending on the portion of the cochlea thatthe aural pressure modulating agent formulation is targeted. Isotonicformulations are provided by the addition of a tonicity agent. Suitabletonicity agents include, but are not limited to any pharmaceuticallyacceptable sugar, salt or any combinations or mixtures thereof, such as,but not limited to dextrose and sodium chloride. In further embodiments,the tonicity agents are present in an amount from about 100 mOsm/kg toabout 500 mOsm/kg. In some embodiments, the tonicity agent is present inan amount from about 200 mOsm/kg to about 400 mOsm/kg, from about 280mOsm/kg to about 320 mOsm/kg. The amount of tonicity agents will dependon the target structure of the pharmaceutical formulation, as describedherein.

Useful tonicity compositions also include one or more salts in an amountrequired to bring osmolality of the composition into an acceptable rangefor the perilymph or the endolymph. Such salts include those havingsodium, potassium or ammonium cations and chloride, citrate, ascorbate,borate, phosphate, bicarbonate, sulfate, thiosulfate or bisulfiteanions; suitable salts include sodium chloride, potassium chloride,sodium thiosulfate, sodium bisulfite and ammonium sulfate.

In some embodiments, the auris-acceptable gel formulations disclosedherein alternatively or additionally contains preservatives to preventmicrobial growth. Suitable auris-acceptable preservatives for use in theenhanced viscosity formulations described herein include, but are notlimited to benzoic acid, boric acid, p-hydroxybenzoates, alcohols,quarternary compounds, stabilized chlorine dioxide, mercurials, such asmerfen and thiomersal, mixtures of the foregoing and the like.

In a further embodiment, the preservative is, by way of example only, anantimicrobial agent, within the auris-acceptable formulations presentedherein. In one embodiment, the formulation includes a preservative suchas by way of example only, methyl paraben, sodium bisulfite, sodiumthiosulfate, ascorbate, chorobutanol, thimerosal, parabens, benzylalcohol, phenylethanol and others. In another embodiment, the methylparaben is at a concentration of about 0.05% to about 1.0%, about 0.1%to about 0.2%. In a further embodiment, the gel is prepared by mixingwater, methylparaben, hydroxyethylcellulose and sodium citrate. In afurther embodiment, the gel is prepared by mixing water, methylparaben,hydroxyethylcellulose and sodium acetate. In a further embodiment, themixture is sterilized by autoclaving at 120° C. for about 20 minutes,and tested for pH, methylparaben concentration and viscosity beforemixing with the appropriate amount of the aural pressure modulatingagent disclosed herein.

Suitable auris-acceptable water soluble preservatives which are employedin the drug delivery vehicle include sodium bisulfite, sodiumthiosulfate, ascorbate, chorobutanol, thimerosal, parabens, benzylalcohol, Butylated hydroxytoluene (BHT), phenylethanol and others. Theseagents are present, generally, in amounts of about 0.001% to about 5% byweight and, preferably, in the amount of about 0.01 to about 2% byweight. In some embodiments, auris-compatible formulations describedherein are free of preservatives.

Round window membrane Penetration Enhancers

In another embodiment, the formulation further comprises one or moreround window membrane penetration enhancers. Penetration across theround window membrane is enhanced by the presence of round windowmembrane penetration enhancers. Round window membrane penetrationenhancers are chemical entities that facilitate transport ofcoadministered substances across the round window membrane. Round windowmembrane penetration enhancers are grouped according to chemicalstructure. Surfactants, both ionic and non-ionic, such as sodium laurylsulfate, sodium laurate, polyoxyethylene-20-cetyl ether, laureth-9,sodium dodecylsulfate, dioctyl sodium sulfosuccinate,polyoxyethylene-9-lauryl ether (PLE), Tween® 80,nonylphenoxypolyethylene (NP-POE), polysorbates and the like, functionas round window membrane penetration enhancers. Bile salts (such assodium glycocholate, sodium deoxycholate, sodium taurocholate, sodiumtaurodihydrofusidate, sodium glycodihydrofusidate and the like), fattyacids and derivatives (such as oleic acid, caprylic acid, mono- anddi-glycerides, lauric acids, acylcholines, caprylic acids,acylcarnitines, sodium caprates and the like), chelating agents (such asEDTA, citric acid, salicylates and the like), sulfoxides (such asdimethyl sulfoxide (DMSO), decylmethyl sulfoxide and the like), andalcohols (such as ethanol, isopropanol, glycerol, propanediol and thelike) also function as round window membrane penetration enhancers.

In some embodiments, the auris acceptable penetration enhancer is asurfactant comprising an alkyl-glycoside wherein the alkyl glycoside istetradecyl-β-D-maltoside. In some embodiments, the auris acceptablepenetration enhancer is a surfactant comprising an alkyl-glycosidewherein the alkyl glycoside is dodecyl-maltoside. In certain instances,the penetration enhancing agent is a hyaluronidase. In certaininstances, a hyaluronidase is a human or bovine hyaluronidase. In someinstances, a hyaluronidase is a human hyaluronidase (e.g., hyaluronidasefound in human sperm, PH20 (Halozyme), Hyelenex® (Baxter International,Inc.)). In some instances, a hyaluronidase is a bovine hyaluronidase(e.g., bovine testicular hyaluronidase, Amphadase® (AmphastarPharmaceuticals), Hydase® (PrimaPharm, Inc). In some instances, ahyluronidase is an ovine hyaluronidase, Vitrase® (ISTA Pharmaceuticals).In certain instances, a hyaluronidase described herein is a recombinanthyaluronidase. In some instances, a hyaluronidase described herein is ahumanized recombinant hyaluronidase. In some instances, a hyaluronidasedescribed herein is a pegylated hyaluronidase (e.g., PEGPH20(Halozyme)). In addition, the peptide-like penetration enhancersdescribed in U.S. Pat. Nos. 7,151,191, 6,221,367 and 5,714,167, hereinincorporated by references for such disclosure, are contemplated as anadditional embodiment. These penetration enhancers are amino-acid andpeptide derviatives and enable drug absorption by passive transcellulardiffusion without affecting the integrity of membranes or intercellulartight junctions.

Round window membrane Permeable Liposomes

Liposomes or lipid particles may also be employed to encapsulate theaural pressure modulating agent formulations or compositions.Phospholipids that are gently dispersed in an aqueous medium formmultilayer vesicles with areas of entrapped aqueous media separating thelipid layers. Sonication, or turbulent agitation, of these multilayerveiscles results in the formation of single layer vesicles, commonlyreferred to as liposomes, with sizes of about 10-1000 nm. Theseliposomes have many advantages as aural pressure modulating agents orother pharmaceutical agent carriers. They are biologically inert,biodegradable, non-toxic and non-antigenic. Liposomes are formed invarious sizes and with varying compositions and surface properties.Additionally, they are able to entrap a wide variety of agents andrelease the agent at the site of liposome collapse.

Suitable phospholipids for use in auris-acceptable liposomes here are,for example, phosphatidyl cholines, ethanolamines and serines,sphingomyelins, cardiolipins, plasmalogens, phosphatidic acids andcerebrosides, in particular those which are soluble together with theaural pressure modulating agents herein in non-toxic, pharmaceuticallyacceptable organic solvents. Preferred phospholipids are, for example,phosphatidyl choline, phosphatidyl ethanolmine, phosphatidyl serine,phosphatidyl inositol, lysophosphatidyl choline, phosphatidyl glyceroland the like, and mixtures thereof especially lecithin, e.g. soyalecithin. The amount of phospholipid used in the present formulationrange from about 10 to about 30%, preferably from about 15 to about 25%and in particular is about 20%.

Lipophilic additives may be employed advantageously to modifyselectively the characteristics of the liposomes. Examples of suchadditives include by way of example only, stearylamine, phosphatidicacid, tocopherol, cholesterol, cholesterol hemisuccinate and lanolinextracts. The amount of lipophilic additive used range from 0.5 to 8%,preferably from 1.5 to 4% and in particular is about 2%. Generally, theratio of the amount of lipophilic additive to the amount of phospholipidranges from about 1:8 to about 1:12 and in particular is about 1:10.Said phospholipid, lipophilic additive and the aural pressure modulatingagent and other pharmaceutical compounds are employed in conjunctionwith a non-toxic, pharmaceutically acceptable organic solvent systemwhich dissolve said ingredients. Said solvent system not only mustdissolve the aural pressure modulating agent completely, but it also hasto allow the formulation of stable single bilayered liposomes. Thesolvent system comprises dimethylisosorbide and tetraglycol (glycofurol,tetrahydrofurfuryl alcohol polyethylene glycol ether) in an amount ofabout 8 to about 30%. In said solvent system, the ratio of the amount ofdimethylisosorbide to the amount of tetraglycol range from about 2:1 toabout 1:3, in particular from about 1:1 to about 1:2.5 and preferably isabout 1:2. The amount of tetraglycol in the final composition thus varyfrom 5 to 20%, in particular from 5 to 15% and preferably isapproximately 10%. The amount of dimethylisosorbide in the finalcomposition thus range from 3 to 10%, in particular from 3 to 7% andpreferably is approximately 5%.

The term “organic component” as used hereinafter refers to mixturescomprising said phospholipid, lipophilic additives and organic solvents.The aural pressure modulating agent may be dissolved in the organiccomponent, or other means to maintain full activity of the agent. Theamount of aural pressure modulating agent in the final formulation mayrange from 0.1 to 5.0%. In addition, other ingredients such asanti-oxidants may be added to the organic component. Examples includetocopherol, butylated hydroxyanisole, butylated hydroxytoluene, ascorbylpalmitate, ascorbyl oleate and the like.

Liposomal formulations are alternatively prepared, for aural pressuremodulating agents or other pharmaceutical agents that are moderatelyheat-resistant, by (a) heating the phospholipid and the organic solventsystem to about 60-80° C. in a vessel, dissolving the active ingredient,then adding any additional formulating agents, and stirring the mixtureuntil complete dissolution is obtained; (b) heating the aqueous solutionto 90-95° C. in a second vessel and dissolving the preservativestherein, allowing the mixture to cool and then adding the remainder ofthe auxiliary formulating agents and the remainder of the water, andstirring the mixture until complete dissolution is obtained; thuspreparing the aqueous component; (c) transferring the organic phasedirectly into the aqueous component, while homogenizing the combinationwith a high performance mixing apparatus, for example, a high-shearmixer; and (d) adding a viscosity enhancing agent to the resultingmixture while further homogenizing. The aqueous component is optionallyplaced in a suitable vessel which is equipped with a homogenizer andhomogenization is effected by creating turbulence during the injectionof the organic component. Any mixing means or homogenizer which exertshigh shear forces on the mixture may be employed. Generally, a mixercapable of speeds from about 1,500 to 20,000 rpm, in particular fromabout 3,000 to about 6,000 rpm may be employed. Suitable viscosityenhancing agents for use in process step (d) are for example, xanthangum, hydroxypropyl cellulose, hydroxypropyl methylcellulose or mixturesthereof. The amount of viscosity enhancing agent depends on the natureand the concentration of the other ingredients and in general rangesfrom about 0.5 to 2.0%, or approximately 1.5%. In order to preventdegradation of the materials used during the preparation of theliposomal formulation, it is advantageous to purge all solutions with aninert gas such as nitrogen or argon, and to conduct all steps under aninert atmosphere. Liposomes prepared by the above described methodusually contain most of the active ingredient bound in the lipid bilayerand separation of the liposomes from unencapsulated material is notrequired.

In other embodiments, the auris-acceptable formulations, including gelformulations and viscosity-enhanced formulations, further includeexcipients, other medicinal or pharmaceutical agents, carriers,adjuvants, such as preserving, stabilizing, wetting or emulsifyingagents, solution promoters, salts, solubilizers, an antifoaming agent,an antioxidant, a dispersing agent, a wetting agent, a surfactant, andcombinations thereof.

Suitable carriers for use in an auris-acceptable formulation describedherein include, but are not limited to, any pharmaceutically acceptablesolvent compatible with the targeted auris structure's physiologicalenvironment. In other embodiments, the base is a combination of apharmaceutically acceptable surfactant and solvent.

In some embodiments, other excipients include, sodium stearyl fumarate,diethanolamine cetyl sulfate, isostearate, polyethoxylated castor oil,nonoxyl 10, octoxynol 9, sodium lauryl sulfate, sorbitan esters(sorbitan monolaurate, sorbitan monooleate, sorbitan monopalmitate,sorbitan monostearate, sorbitan sesquioleate, sorbitan trioleate,sorbitan tristearate, sorbitan laurate, sorbitan oleate, sorbitanpalmitate, sorbitan stearate, sorbitan dioleate, sorbitansesqui-isostearate, sorbitan sesquistearate, sorbitan tri-isostearate),lecithin pharmaceutical acceptable salts thereof and combinations ormixtures thereof.

In other embodiments, the carrier is a polysorbate. Polysorbates arenonionic surfactants of sorbitan esters. Polysorbates useful in thepresent disclosure include, but are not limited to polysorbate 20,polysorbate 40, polysorbate 60, polysorbate 80 (Tween 80) and anycombinations or mixtures thereof. In further embodiments, polysorbate 80is utilized as the pharmaceutically acceptable carrier.

In one embodiment, water-soluble glycerin-based auris-acceptableenhanced viscosity formulations utilized in the preparation ofpharmaceutical delivery vehicles comprise at least one aural pressuremodulating agent containing at least about 0.1% of the water-solubleglycerin compound or more. In some embodiments, the percentage of auralpressure modulating agent is varied between about 1% and about 95%,between about 5% and about 80%, between about 10% and about 60% or moreof the weight or volume of the total pharmaceutical formulation. In someembodiments, the amount of the compound(s) in each therapeuticallyuseful aural pressure modulating agent formulation is prepared in such away that a suitable dosage will be obtained in any given unit dose ofthe compound. Factors such as solubility, bioavailability, biologicalhalf-life, route of administration, product shelf life, as well as otherpharmacological considerations are contemplated herein.

If desired, the auris-acceptable pharmaceutical gels also containco-solvents, preservatives, cosolvents, ionic strength and osmolalityadjustors and other excipients in addition to buffering agents. Suitableauris-acceptable water soluble buffering agents are alkali or alkalineearth metal carbonates, phosphates, bicarbonates, citrates, borates,acetates, succinates and the like, such as sodium phosphate, citrate,borate, acetate, bicarbonate, carbonate and tromethamine (TRIS). Theseagents are present in amounts sufficient to maintain the pH of thesystem at 7.4±0.2 and preferably, 7.4. As such, the buffering agent isas much as 5% on a weight basis of the total composition.

Cosolvents are used to enhance aural pressure modulating agentsolubility, however, some aural pressure modulating agents or otherpharmaceutical compounds are insoluble. These are often suspended in thepolymer vehicle with the aid of suitable suspending or viscosityenhancing agents.

Moreover, some pharmaceutical excipients, diluents or carriers arepotentially ototoxic. For example, benzalkonium chloride, a commonpreservative, is ototoxic and therefore potentially harmful ifintroduced into the vestibular or cochlear structures. In formulating acontrolled release aural pressure modulating agent formulation, it isadvised to avoid or combine the appropriate excipients, diluents orcarriers to lessen or eliminate potential ototoxic components from theformulation, or to decrease the amount of such excipients, diluents orcarriers. Optionally, a controlled release aural pressure modulatingagent formulation includes otoprotective agents, such as antioxidants,alpha lipoic acid, calcium, fosfomycin or iron chelators, to counteractpotential ototoxic effects that may arise from the use of specifictherapeutic agents or excipients, diluents or carriers.

Thof therapeutically acceptable otic formulations:

Example Formulation Example Characteristics Chitosan glycerophosphatetunable degradation of matrix in vitro (CGP) tunable TACE inhibitorrelease in vitro: e.g., ~50% of drug released after 24 hrs biodegradablecompatible with drug delivery to the inner ear suitable formacromolecules and hydrophobic drugs PEG-PLGA-PEG triblock tunable highstability: e.g., maintains mechanical integrity >1 polymers month invitro tunable fast release of hydrophilic drugs: e.g., ~50% of drugreleased after 24 hrs, and remainder released over ~5 days tunable slowrelease of hydrophobic drugs: e.g., ~80% released after 8 weeksbiodegradable subcutaneous injection of solution: e.g., gel forms withinseconds and is intact after 1 month PEO-PPO-PEO triblock Tunable sol-geltransition temperature: e.g., decreases with copolymers (e.g., Pluronicincreasing F127 concentration or Poloxameres) (e.g., F127) Chitosanglycerophosphate CGP formulation tolerates liposomes: e.g., up to 15uM/ml with drug-loaded liposomes liposomes. liposomes tunably reducedrug release time (e.g., up to 2 weeks in vitro). increase in liposomediameter optionally reduces drug release kinetics (e.g., liposome sizebetween 100 and 300 nm) release parameters are controlled by changingcomposition of liposomes

The formulations disclosed herein alternatively encompass anotoprotectant agent in addition to the at least one active agent and/orexcipients, including but not limited to such agents as antioxidants,alpha lipoic acid, calcium, fosfomycin or iron chelators, to counteractpotential ototoxic effects that may arise from the use of specifictherapeutic agents or excipients, diluents or carriers.

Modes of Treatment

Dosing Methods and Schedules

Drugs delivered to the inner ear have been administered systemically viaoral, intravenous or intramuscular routes. However, systemicadministration for pathologies local to the inner ear increases thelikelihood of systemic toxicities and adverse side effects and creates anon-productive distribution of drug in which high levels of drug arefound in the serum and correspondingly lower levels are found at theinner ear.

Intratympanic injection of therapeutic agents is the technique ofinjecting a therapeutic agent behind the tympanic membrane into themiddle and/or inner ear. In one embodiment, the formulations describedherein are administered directly onto the round window membrane viatranstympanic injection. In another embodiment, the aural pressuremodulating agent auris-acceptable formulations described herein areadministered onto the round window membrane via a non-transtympanicapproach to the inner ear. In additional embodiments, the formulationdescribed herein is administered onto the round window membrane via asurgical approach to the round window membrane comprising modificationof the crista fenestrae cochleae.

In one embodiment the delivery system is a syringe and needle apparatusthat is capable of piercing the tympanic membrane and directly accessingthe round window membrane or crista fenestrae cochleae of the aurisinterna. In some embodiments, the needle on the syringe is wider than a18 gauge needle. In another embodiment, the needle gauge is from 18gauge to 31 gauge. In a further embodiment, the needle gauge is from 25gauge to 30 gauge. Depending upon the thickness or viscosity of theaural pressure modulating agent compositions or formulations, the gaugelevel of the syringe or hypodermic needle may be varied accordingly. Inanother embodiment, the internal diameter of the needle can be increasedby reducing the wall thickness of the needle (commonly referred as thinwall or extra thin wall needles) to reduce the possibility of needleclogging while maintaining an adequate needle gauge.

In another embodiment, the needle is a hypodermic needle used forinstant delivery of the gel formulation. The hypodermic needle may be asingle use needle or a disposable needle. In some embodiments, a syringemay be used for delivery of the pharmaceutically acceptable gel-basedaural pressure modulating agent-containing compositions as disclosedherein wherein the syringe has a press-fit (Luer) or twist-on(Luer-lock) fitting. In one embodiment, the syringe is a hypodermicsyringe. In another embodiment, the syringe is made of plastic or glass.In yet another embodiment, the hypodermic syringe is a single usesyringe. In a further embodiment, the glass syringe is capable of beingsterilized. In yet a further embodiment, the sterilization occursthrough an autoclave. In another embodiment, the syringe comprises acylindrical syringe body wherein the gel formulation is stored beforeuse. In other embodiments, the syringe comprises a cylindrical syringebody wherein the aural pressure modulating agent pharmaceuticallyacceptable gel-based compositions as disclosed herein is stored beforeuse which conveniently allows for mixing with a suitablepharmaceutically acceptable buffer. In other embodiments, the syringemay contain other excipients, stabilizers, suspending agents, diluentsor a combination thereof to stabilize or otherwise stably store theaural pressure modulating agent or other pharmaceutical compoundscontained therein.

In some embodiments, the syringe comprises a cylindrical syringe bodywherein the body is compartmentalized in that each compartment is ableto store at least one component of the auris-acceptable aural pressuremodulating agent gel formulation. In a further embodiment, the syringehaving a compartmentalized body allows for mixing of the componentsprior to injection into the auris media or auris interna. In otherembodiments, the delivery system comprises multiple syringes, eachsyringe of the multiple syringes contains at least one component of thegel formulation such that each component is pre-mixed prior to injectionor is mixed subsequent to injection. In a further embodiment, thesyringes disclosed herein comprise at least one reservoir wherein the atleast one reservoir comprises an aural pressure modulating agent, or apharmaceutically acceptable buffer, or a viscosity enhancing agent, suchas a gelling agent or a combination thereof. Commercially availableinjection devices are optionally employed in their simplest form asready-to-use plastic syringes with a syringe barrel, needle assemblywith a needle, plunger with a plunger rod, and holding flange, toperform an intratympanic injection.

In some embodiments, the delivery device is an apparatus designed foradministration of therapeutic agents to the middle and/or inner ear. Byway of example only: GYRUS Medical Gmbh offers micro-otoscopes forvisualization of and drug delivery to the round window niche; Arenberghas described a medical treatment device to deliver fluids to inner earstructures in U.S. Pat. Nos. 5,421,818; 5,474,529; and 5,476,446, eachof which is incorporated by reference herein for such disclosure. U.S.patent application Ser. No. 08/874,208, which is incorporated herein byreference for such disclosure, describes a surgical method forimplanting a fluid transfer conduit to deliver therapeutic agents to theinner ear. U.S. Patent Application Publication 2007/0167918, which isincorporated herein by reference for such disclosure, further describesa combined otic aspirator and medication dispenser for intratympanicfluid sampling and medicament application.

The formulations described herein, and modes of administration thereof,are also applicable to methods of direct instillation or perfusion ofthe inner ear compartments. Thus, the formulations described herein areuseful in surgical procedures including, by way of non-limitingexamples, cochleostomy, labyrinthotomy, mastoidectomy, stapedectomy,endolymphatic sacculotomy or the like.

The auris-acceptable compositions or formulations containing the auralpressure modulating agent compound(s) described herein are administeredfor prophylactic and/or therapeutic treatments. In therapeuticapplications, the aural pressure modulating agent compositions areadministered to a patient already suffering from an autoimmune disease,condition or disorder, in an amount sufficient to cure or at leastpartially arrest the symptoms of the disease, disorder or condition.Amounts effective for this use will depend on the severity and course ofthe disease, disorder or condition, previous therapy, the patient'shealth status and response to the drugs, and the judgment of thetreating physician.

In the case wherein the patient's condition does not improve, upon thedoctor's discretion the administration of the aural pressure modulatingagent compounds may be administered chronically, that is, for anextended period of time, including throughout the duration of thepatient's life in order to ameliorate or otherwise control or limit thesymptoms of the patient's disease or condition.

In the case wherein the patient's status does improve, upon the doctor'sdiscretion the administration of the aural pressure modulating agentcompounds may be given continuously; alternatively, the dose of drugbeing administered may be temporarily reduced or temporarily suspendedfor a certain length of time (i.e., a “drug holiday”). The length of thedrug holiday varies between 2 days and 1 year, including by way ofexample only, 2 days, 3 days, 4 days, 5 days, 6 days, 7 days, 10 days,12 days, 15 days, 20 days, 28 days, 35 days, 50 days, 70 days, 100 days,120 days, 150 days, 180 days, 200 days, 250 days, 280 days, 300 days,320 days, 350 days, and 365 days. The dose reduction during a drugholiday may be from 10%-100%, including by way of example only 10%, 15%,20%, 25%, 30%, 35%, 40%, 45%, 50%, 55%, 60%, 65%, 70%, 75%, 80%, 85%,90%, 95%, and 100%.

Once improvement of the patient's otic conditions has occurred, amaintenance aural pressure modulating agent dose is administered ifnecessary. Subsequently, the dosage or the frequency of administration,or both, is optionally reduced, as a function of the symptoms, to alevel at which the improved disease, disorder or condition is retained.In certain embodiments, patients require intermittent treatment on along-term basis upon any recurrence of symptoms.

The amount of aural pressure modulating agent that will correspond tosuch an amount will vary depending upon factors such as the particularcompound, disease condition and its severity, according to theparticular circumstances surrounding the case, including, e.g., thespecific aural pressure modulating agent being administered, the routeof administration, the autoimmune condition being treated, the targetarea being treated, and the subject or host being treated. In general,however, doses employed for adult human treatment will typically be inthe range of 0.02-50 mg per administration, preferably 1-15 mg peradministration. The desired dose is presented in a single dose or asdivided doses administered simultaneously (or over a short period oftime) or at appropriate intervals.

In some embodiments, the initial administration is a particular auralpressure modulating agent and the subsequent administration a differentformulation or aural pressure modulating agent.

Pharmacokinetics of Controlled Release Formulations

In one embodiment, the formulations disclosed herein additionallyprovides an immediate release of an aural pressure modulating agent fromthe composition, or within 1 minute, or within 5 minutes, or within 10minutes, or within 15 minutes, or within 30 minutes, or within 60minutes or within 90 minutes. In other embodiments, a therapeuticallyeffective amount of at least one aural pressure modulating agent isreleased from the composition immediately, or within 1 minute, or within5 minutes, or within 10 minutes, or within 15 minutes, or within 30minutes, or within 60 minutes or within 90 minutes. In certainembodiments the composition comprises an auris-pharmaceuticallyacceptable gel formulation providing immediate release of at least oneaural pressure modulating agent. Additional embodiments of theformulation may also include an agent that enhances the viscosity of theformulations included herein.

In other or further embodiments, the formulation provides an extendedrelease formulation of at least one aural pressure modulating agent. Incertain embodiments, diffusion of at least one aural pressure modulatingagent from the formulation occurs for a time period exceeding 5 minutes,or 15 minutes, or 30 minutes, or 1 hour, or 4 hours, or 6 hours, or 12hours, or 18 hours, or 1 day, or 2 days, or 3 days, or 4 days, or 5days, or 6 days, or 7 days, or 10 days, or 12 days, or 14 days, or 18days, or 21 days, or 25 days, or 30 days, or 45 days, or 2 months or 3months or 4 months or 5 months or 6 months or 9 months or 1 year. Inother embodiments, a therapeutically effective amount of at least oneaural pressure modulating agent is released from the formulation for atime period exceeding 5 minutes, or 15 minutes, or 30 minutes, or 1hour, or 4 hours, or 6 hours, or 12 hours, or 18 hours, or 1 day, or 2days, or 3 days, or 4 days, or 5 days, or 6 days, or 7 days, or 10 days,or 12 days, or 14 days, or 18 days, or 21 days, or 25 days, or 30 days,or 45 days, or 2 months or 3 months or 4 months or 5 months or 6 monthsor 9 months or 1 year.

In other embodiments, the formulation provides both an immediate releaseand an extended release formulation of an aural pressure modulatingagent. In yet other embodiments, the formulation contains a 0.25:1ratio, or a 0.5:1 ratio, or a 1:1 ratio, or a 1:2 ratio, or a 1:3, or a1:4 ratio, or a 1:5 ratio, or a 1:7 ratio, or a 1:10 ratio, or a 1:15ratio, or a 1:20 ratio of immediate release and extended releaseformulations. In a further embodiment the formulation provides animmediate release of a first aural pressure modulating agent and anextended release of a second aural pressure modulating agent or othertherapeutic agent. In yet other embodiments, the formulation provides animmediate release and extended release formulation of at least one auralpressure modulating agent, and at least one therapeutic agent. In someembodiments, the formulation provides a 0.25:1 ratio, or a 0.5:1 ratio,or a 1:1 ratio, or a 1:2 ratio, or a 1:3, or a 1:4 ratio, or a 1:5ratio, or a 1:7 ratio, or a 1:10 ratio, or a 1:15 ratio, or a 1:20 ratioof immediate release and extended release formulations of a first auralpressure modulating agent and second therapeutic agent, respectively.

In a specific embodiment the formulation provides a therapeuticallyeffective amount of at least one aural pressure modulating agent at thesite of disease with essentially no systemic exposure. In an additionalembodiment the formulation provides a therapeutically effective amountof at least one aural pressure modulating agent at the site of diseasewith essentially no detectable systemic exposure. In other embodiments,the formulation provides a therapeutically effective amount of at leastone aural pressure modulating agent at the site of disease with littleor no detectable systemic exposure.

The combination of immediate release, delayed release and/or extendedrelease aural pressure modulating agent compositions or formulations maybe combined with other pharmaceutical agents, as well as the excipients,diluents, stabilizers, tonicity agents and other components disclosedherein. As such, depending upon the aural pressure modulating agentused, the thickness or viscosity desired, or the mode of deliverychosen, alternative aspects of the embodiments disclosed herein arecombined with the immediate release, delayed release and/or extendedrelease embodiments accordingly.

In certain embodiments, the pharmacokinetics of the aural pressuremodulating agent formulations described herein are determined byinjecting the formulation on or near the round window membrane of a testanimal (including by way of example, a guinea pig or a chinchilla). At adetermined period of time (e.g., 6 hours, 12 hours, 1 day, 2 days, 3days, 4 days, 5 days, 6 days, and 7 days for testing thepharmacokinetics of a formulation over a 1 week period), the test animalis euthanized and a 5 mL sample of the perilymph fluid is tested. Theinner ear removed and tested for the presence of the aural pressuremodulating agent. As needed, the level of aural pressure modulatingagent is measured in other organs. In addition, the systemic level ofthe aural pressure modulating agent is measured by withdrawing a bloodsample from the test animal. In order to determine whether theformulation impedes hearing, the hearing of the test animal isoptionally tested.

Alternatively, an inner ear is provided (as removed from a test animal)and the migration of the aural pressure modulating agent is measured. Asyet another alternative, an in vitro model of a round window membrane isprovided and the migration of the aural pressure modulating agent ismeasured.

Kits/Articles of Manufacture

The disclosure also provides kits for preventing, treating orameliorating the symptoms of a disease or disorder in a mammal. Suchkits generally will comprise one or more of the aural pressuremodulating agent controlled-release compositions or devices disclosedherein, and instructions for using the kit. The disclosure alsocontemplates the use of one or more of the aural pressure modulatingagent controlled-release compositions, in the manufacture of medicamentsfor treating, abating, reducing, or ameliorating the symptoms of adisease, dysfunction, or disorder in a mammal, such as a human that has,is suspected of having, or at risk for developing an inner ear disorder.

In some embodiments, kits include a carrier, package, or container thatis compartmentalized to receive one or more containers such as vials,tubes, and the like, each of the container(s) including one of theseparate elements to be used in a method described herein. Suitablecontainers include, for example, bottles, vials, syringes, and testtubes. In other embodiments, the containers are formed from a variety ofmaterials such as glass or plastic.

The articles of manufacture provided herein contain packaging materials.Packaging materials for use in packaging pharmaceutical products arealso presented herein. See, e.g., U.S. Pat. Nos. 5,323,907, 5,052,558and 5,033,252. Examples of pharmaceutical packaging materials include,but are not limited to, blister packs, bottles, tubes, inhalers, pumps,bags, vials, containers, syringes, bottles, and any packaging materialsuitable for a selected formulation and intended mode of administrationand treatment. A wide array of aural pressure modulating agentformulations compositions provided herein are contemplated as are avariety of treatments for any disease, disorder, or condition that wouldbenefit by controlled release administration of a aural pressuremodulating agent to the inner ear.

In some embodiments, a kit includes one or more additional containers,each with one or more of various materials (such as reagents, optionallyin concentrated form, and/or devices) desirable from a commercial anduser standpoint for use of a formulation described herein. Non-limitingexamples of such materials include, but not limited to, buffers,diluents, filters, needles, syringes; carrier, package, container, vialand/or tube labels listing contents and/or instructions for use andpackage inserts with instructions for use. A set of instructions isoptionally included. In a further embodiment, a label is on orassociated with the container. In yet a further embodiment, a label ison a container when letters, numbers or other characters forming thelabel are attached, molded or etched into the container itself; a labelis associated with a container when it is present within a receptacle orcarrier that also holds the container, e.g., as a package insert. Inother embodiments a label is used to indicate that the contents are tobe used for a specific therapeutic application. In yet anotherembodiment, a label also indicates directions for use of the contents,such as in the methods described herein.

In certain embodiments, the pharmaceutical compositions are presented ina pack or dispenser device which contains one or more unit dosage formscontaining a compound provided herein. In another embodiment, the packfor example contains metal or plastic foil, such as a blister pack. In afurther embodiment, the pack or dispenser device is accompanied byinstructions for administration. In yet a further embodiment, the packor dispenser is also accompanied with a notice associated with thecontainer in form prescribed by a governmental agency regulating themanufacture, use, or sale of pharmaceuticals, which notice is reflectiveof approval by the agency of the form of the drug for human orveterinary administration. In another embodiment, such notice, forexample, is the labeling approved by the U.S. Food and DrugAdministration for prescription drugs, or the approved product insert.In yet another embodiment, compositions containing a compound providedherein formulated in a compatible pharmaceutical carrier are alsoprepared, placed in an appropriate container, and labeled for treatmentof an indicated condition.

EXAMPLES Example 1 Preparation of a Thermoreversible Gel VP2 AntagonistFormulation

Quantity Ingredient (mg/g of formulation) Lixivaptan 20.0 methylparaben1.0 HPMC 10.0 Poloxamer 407 180.0 TRIS HCl buffer (0.1 M) 789.0

A 10-g batch of gel formulation containing 2.0% of lixivaptan isprepared by suspending 1.80 g of Poloxamer 407 (BASF Corp.) in 5.00 g ofTRIS HCl buffer (0.1 M) and the components are mixed under agitationovernight at 4° C. to ensure complete dissolution. The lixivaptan (200.0mg), hydroxypropylmethylcellulose (100.0 mg), methylparaben (10 mg) andadditional TRIS HCl buffer (0.1 M) (2.89 g) is added and furtherstirring allowed until complete dissolution is observed. The mixture ismaintained below room temperature until use.

Example 2 Preparation of Liposomal VP2 Antagonist Formation

Quantity Ingredient (mg/g of cream) Lixivaptan 5.0 soya lecithin 200.0cholesterol 20.0 tetraglycol 100.0 dimethylisosorbide 50.0 methylparaben2.0 propylparaben 0.2 BHT 0.1 sodium chloride 1.0 HPMC 15.0 sodiumhydroxide 0.6 citric acid 1.0 purified water, USP 603.6

Heat the soya lecithin, tetraglycol and dimethyl isosorbide to about70-75° C. Dissolve the lixivaptan, cholesterol and butylatedhydroxytoluene in the heated mixture. Stir until complete dissolution isobtained. Heat about one third of the water to 80-95° C. in a separatevessel and dissolve the preservatives methylparaben and propylparaben inthe heated water while stirring. Allow the solution to cool to about 25°C. and then add the disodium edetate, sodium chloride, sodium hydroxideand citric acid. Add the remainder of the water and stir to obtain acomplete solution. Transfer the organic mixture into the aqueous mixtureby means of a vacuum, while homogenizing the combination with ahigh-shear mixer until a homogeneous product is obtained. Add thehydroxypropyl methylcellulose into the biphasic mixture by means of avacuum while homogenizing with a mixer. The homogenizer is a Silversonhigh-shear mixer operating at approximately 3000 rpm. Single bilayeredliposomes are formed. The white lipogel cream is ready for use.

Example 3 Preparation of a VP2 Antagonist Nanoparticle Formulation

750 mg (15 mg/ml theoretical) of a diblock copolymer consisting of thecombination of a poly(d,l-lactic acid) of mass 30 kD and of apolyethylene glycol of mass 2 kD (PLA-PEG) and 250 mg (5 mg/mltheoretical) of tolvaptan is dissolved in 20 ml of ethyl acetate(solution A). 175 mg of lecithin E80 and 90 mg of sodium oleate isdispersed in 50 ml of 5% w/v glucose solution (solution B). Solution Ais emulsified in solution B with an Ultra-turrax stirrer and thepre-emulsion is then introduced into a Microfluidizer 110 S.RTM. typehomogenizer for 10 minutes at 10° C. The volume of emulsion recovered isabout 70 ml (70 g). The ethyl acetate is removed using a rotaryevaporator at reduced pressure (100 mm of mercury) to a suspensionvolume of about 45 ml (45 g).

Example 4 Preparation of a 5% Cyclodextrin VP2 Antagonist Formulation

To a suitable 150 mL glass vessel is added tolvaptan (5.0 g), sterile 2%dibasic sodium phosphate dodecahydrate solution (9.0 g) andHydroxypropyl-®-Cyclodextrin (50 g). The resulting mixture is stirreduntil a clear solution is formed. To this solution is added sterile 2%polysorbate 80 solution (5 g), sterile 2% stock HPMC 2910 (E4M) solution(2.5 g) and 5% sterile sodium chloride solution (11 g), and stirring iscontinued until homogeneous. Sterile water for injection is added to getto 95% of batch size. The solution is stirred at rt for 30 min and pH isadjusted to 7.2. Finally, water for injection is added to get a finalbatch size of 100 g.

Example 5 Preparation of a Mucoadhesive Thermoreversible Gel VP2Antagonist Formulation

Quantity Ingredient (mg/g of formulation) SR-121463 20.0 methylparaben1.0 HPMC 10.0 Carbopol 934P 2.0 Poloxamer 407 180.0 TRIS HCl buffer (0.1M) 787.0

A 10-g batch of mucoadhesive, gel formulation containing 2.0% ofSR-121463 is prepared by suspending 2.0 mg of Carbopol 934P and 1.80 gof Poloxamer 407 (BASF Corp.) in 5.00 g of TRIS HCl buffer (0.1 M) andthe components are mixed under agitation overnight at 4° C. to ensurecomplete dissolution. The SR-121463 (200.0 mg),hydroxypropylmethylcellulose (100.0 mg), methylparaben (10 mg) andadditional TRIS HCl buffer (0.1 M) (2.87 g) are added and furtherstirring allowed until complete dissolution is observed. The mixture ismaintained below room temperature until use.

Example 6 Preparation of a 50% VP2 Antagonist 95:5 d,l-PLGA MicrosphereFormulation

Twenty-five grams (25 g) of 95:5 d,l-PLGA and 25 g of OPC-31260 arecodissolved in 196 g ethyl acetate in an Erlenmeyer flask at 52° C. Thedrug/polymer solution is added to a 1000 ml glass jacketed reactorcontaining 550 g of 5% aqueous polyvinyl alcohol containing 9.7 g ofethyl acetate. Reactor contents are stirred with an overhead stir motorand the temperature is maintained at 52° C. by a circulating bath. Theemulsion size is monitored by light microscopy and the stirring isstopped when the particle size is found to be in the desired size range(less than 300 microns), usually after about 2 minutes. The stir speedis reduced to avoid further size reduction of the sterilized emulsion.After stirring for a total of 4 minutes, the reactor contents arepressure-transferred into 40 liters of water at 12° C. After stirringfor 20 minutes, the hardened microspheres are isolated and the productthen transferred into 20 liters of water at 12° C. After approximately 3hours, the second wash is transferred onto a sieve stack composed of 25,45, 90, 150, and 212 micron openings. The product on the sieves iswashed with copious amounts of cold water to separate the differentsizes of microspheres. After drying on the sieves overnight, thedifferent fractions are collected and drying was continued under vacuumat room temperature. Formulations with other drug levels are prepared bysimply adjusting the polymer/drug ratio.

Example 7 Preparation of a 50% VP2 Antagonist 65:35 d,l-PLGA MicrosphereFormulation

Microspheres were produced by the method of Example 6 except that adifferent biodegradable polymer matrix was utilized. A 65:35 d,l-PLGApolymer was used in place of the 95:5 polymer indicated in Example 6.

Example 8 Preparation of a Mucoadhesive Cyclodextrin-based VP2Antagonist Formulation

Quantity Ingredient (mg/g of formulation) Lixivaptan 20.0 HP ® CD 500propylene glycol 50 paraffin oil 200 trihydroxystearate 10 cetyldimethicon copolyol 30 water qs ad 1000 phosphate buffer pH 7.4 qs pH7.4

The cream-type formulation is prepared by solubilizing lixivaptan withpropylene glycol and this solution is added to a suspension of HP®CD inwater. A second system is prepared by mixing paraffin oil,trihydroxystearate and cetyl dimethicon copolyol with warming to 60° C.Upon cooling to room temperature, the lipid system is mixed with theaqueous phase in a homogenizer for 30 minutes.

Example 9 Preparation of a Cyclodextrin-containing Thermoreversible Gel2.5% VP2 Antagonist Formulation

Quantity Ingredient (mg/g of formulation) 5% CD solution 500.0methylparaben 1.0 Poloxamer 407 180.0 TRIS HCl buffer (0.1 M) 317.0

The Poloxamer 407 (BASF Corp.) is suspended in the TRIS HCl buffer (0.1M) and the components are mixed under agitation overnight at 4° C. toensure complete dissolution. The cyclodextrin solution from Example 4and methylparaben is added and further stirring allowed until completedissolution is observed. The mixture is maintained below roomtemperature until use.

Example 10 Preparation of a Cyclodextrin-containing MucoadhesiveThermoreversible Gel VP2 Antagonist Formulation

Quantity Ingredient (mg/g of formulation) 5% CD solution 500.0methylparaben 1.0 Poloxamer 407 180.0 Carbopol 934P 2.0 TRIS HCl buffer(0.1 M) 317.0

The Carbopol 934P and Poloxamer 407 (BASF Corp.) is suspended in theTRIS HCl buffer (0.1 M) and the components are mixed under agitationovernight at 4° C. to ensure complete dissolution. The cyclodextrinsolution from Example 4 and methylparaben is added and further stirringallowed until complete dissolution is observed. The mixture ismaintained below room temperature until use.

Example 11 Preparation of a Gel VP2 Antagonist Formulation

Quantity Ingredient (mg/g of formulation) SR-121463 20.0 chitosan 20.0Glycerophosphate disodium 80.0 water 880

A 5 ml solution of acetic acid is titrated to a pH of about 4.0. Thechitosan is added to achieve a pH of about 5.5. The SR-121463 is thendissolved in the chitosan solution. This solution is sterilized byfiltration. A 5 ml aqueous solution of glycerophosphate disodium is alsoprepared and sterilized. The two solutions are mixed and within 2 h at37° C., the desired gel is formed.

Example 12 Preparation of a Gel/Liposome VP2 Antagonist Formulation

Ingredient Quantity SR-121463 20.0 mg/g Liposomes 15 umol/mlChitosan-Glycerophosphate 100.0 mg/g

The liposomes are prepared in the presence of the VP2 antagonistSR-121463 by the reversed-phase evaporation method, where lipids inchloroform or chloroform-methanol (2:1, v/v) are deposited on the sidesof a tube by evaporation of the organic solvent. The lipid film isredissolved in diethyl ether and the aqueous phase (pH 7.4 300 mOsm/kg)containing 20 mM Hepes and 144 mM NaCl is added. The mixture issonicated to obtain a homogeneous emulsion, and then the organic solventis removed under vacuum. The preparation is extruded to obtain therequired liposome size and free components removed by size-exclusionchromatography using a Sephadex G-50 column (Amersham Pharmacia Biotech,Uppsala, Sweden).

To prepare the chitosan-glycerophosphate formulation, a 5 ml solution ofacetic acid is titrated to a pH of about 4.0. The chitosan is added toachieve a pH of about 5.5. This solution is sterilized by filtration. A5 ml aqueous solution of glycerophosphate disodium is also prepared andsterilized. The two solutions are mixed and within 2 h at 37° C., andthe desired gel is formed. The chitosan-glycerophosphate solution isgently mixed with the liposomes at room temperature.

Example 13 Effect of pH on degradation products for autoclaved 17%poloxamer 407NF/2% otic agent in PBS buffer

A stock solution of a 17% poloxamer 407/2% otic agent is prepared bydissolving 351.4 mg of sodium chloride (Fisher Scientific), 302.1 mg ofsodium phosphate dibasic anhydrous (Fisher Scientific), 122.1 mg ofsodium phosphate monobasic anhydrous (Fisher Scientific) and anappropriate amount of an otic agent with 79.3 g of sterile filtered DIwater. The solution is cooled down in a ice chilled water bath and then17.05 g of poloxamer 407NF (SPECTRUM CHEMICALS) is sprinkled into thecold solution while mixing. The mixture is further mixed until thepoloxamer is completely dissolved. The pH for this solution is measured.

17% poloxamer 407/2% otic agent in PBS pH of 5.3. Take an aliquot(approximately 30 mL) of the above solution and adjust the pH to 5.3 bythe addition of 1 M HCl.

17% poloxamer 407/2% otic agent in PBS pH of 8.0. Take an aliquot(approximately 30 mL) of the above stock solution and adjust the pH to8.0 by the addition of 1 M NaOH.

A PBS buffer (pH 7.3) is prepared by dissolving 805.5 mg of sodiumchloride (Fisher Scientific), 606 mg of sodium phosphate dibasicanhydrous (Fisher Scientific), 247 mg of sodium phosphate monobasicanhydrous (Fisher Scientific), then QS to 200 g with sterile filtered DIwater.

A 2% solution of an otic agent in PBS pH 7.3 is prepared by dissolvingan appropriate amount of the otic agent in the PBS buffer and QS to 10 gwith PBS buffer.

One mL samples are individually placed in 3 mL screw cap glass vials(with rubber lining) and closed tightly. The vials are placed in aMarket Forge-sterilmatic autoclave (settings, slow liquids) andsterilized at 250° F. for 15 minutes. After the autoclave the samplesare left to cool down to room temperature and then placed inrefrigerator. The samples are homogenized by mixing the vials whilecold.

Appearance (e.g., discoloration and/or precipitation) is observed andrecorded. HPLC analysis is performed using an Agilent 1200 equipped witha Luna C18(2) 3 μm, 100 Å, 250×4.6 mm column) using a 30-80 acetonitrilegradient (1-10 min) of (water acetonitrile mixture containing 0.05%TFA), for a total run of 15 minutes. Samples are diluted by taking 30 μLof sample and dissolved with 1.5 mL of a 1:1 acetonitrile water mixture.Purity of the otic agent in the autoclaved samples is recorded.

In general the formulation should not have any individual impurity(e.g., degradation product of otic agent) of more than 2% and morepreferably not more than one percent. In addition, the formulationshould not precipitate during storage or change in color aftermanufacturing and storage.

Formulations comprising conivaptan, SR-121463 or micronized lixivaptan,prepared according to the procedure above, are tested using the aboveprocedure to determine the effect of pH on degradation during theautoclaving step.

Example 14 Effect of autoclaving on the release profile and viscosity ofa 17% poloxamer 407NF/2% otic agent in PBS

An aliquot of a sample (autoclaved and not autoclaved) is evaluated forrelease profile and viscosity measurement to evaluate the impact of heatsterilization on the properties of the gel.

Dissolution is performed at 37° C. in snapwells (6.5 mm diameterpolycarbonate membrane with a pore size of 0.4 μm). 0.2 mL of gel isplaced into snapwell and left to harden, then 0.5 mL is placed intoreservoir and shaken using a Labline orbit shaker at 70 rpm. Samples aretaken every hour (0.1 mL withdrawn and replace with warm buffer).Samples are analyzed for poloxamer concentration by UV at 624 nm usingthe cobalt thiocyanate method, against an external calibration standardcurve. In brief, 20 μL of the sample is mixed with 1980 μL of a 15 mMcobalt thiocyanate solution and absorbance measured at 625 nm, using aEvolution 160 UV/Vis spectrophotometer (Thermo Scientific).

The released otic agent is fitted to the Korsmeyer-Peppas equation

$\frac{Q}{Q_{\alpha}} = {{k\; t^{n}} + b}$where Q is the amount of otic agent released at time t, Q_(α) is theoverall released amount of otic agent, k is a release constant of thenth order, n is a dimensionless number related to the dissolutionmechanism and b is the axis intercept, characterizing the initial burstrelease mechanism wherein n=1 characterizes an erosion controlledmechanism. The mean dissolution time (MDT) is the sum of differentperiods of time the drug molecules stay in the matrix before release,divided by the total number of molecules and is calculated by:

${M\; D\; T} = \frac{n\; k^{{- 1}/n}}{n + 1}$

Viscosity measurements are performed using a Brookfield viscometerRVDV-II+P with a CPE-51 spindle rotated at 0.08 rpm (shear rate of 0.31s⁻¹), equipped with a water jacketed temperature control unit(temperature ramped from 15-34° C. at 1.6° C./min). Tgel is defined asthe inflection point of the curve where the increase in viscosity occursdue to the sol-gel transition.

Formulations comprising conivaptan, SR-121463 or micronized lixivaptanprepared according to the procedures described above, are tested usingthe procedure described above to determine Tgel.

Example 15 Effect of addition of a secondary polymer on the degradationproducts and viscosity of a formulation containing 2% otic agent and 17%poloxamer 407NF after heat sterilization (autoclaving)

Solution A. A solution of pH 7.0 comprising sodiumcarboxymethylcellulose (CMC) in PBS buffer is prepared by dissolving178.35 mg of sodium chloride (Fisher Scientific), 300.5 mg of sodiumphosphate dibasic anhydrous (Fisher Scientific), 126.6 mg of sodiumphosphate monobasic anhydrous (Fisher Scientific) dissolved with 78.4 ofsterile filtered DI water, then 1 g of Blanose 7M65 CMC (Hercules,viscosity of 5450 cP @2%) is sprinkled into the buffer solution andheated to aid dissolution, and the solution is then cooled down.

A solution of pH 7.0 comprising 17% poloxamer 407NF/1% CMC/2% otic agentin PBS buffer is made by cooling down 8.1 g of solution A in a icechilled water bath and then adding an appropriate amount of an oticagent followed by mixing. 1.74 g of poloxamer 407NF (Spectrum Chemicals)is sprinkled into the cold solution while mixing. The mixture is furthermixed until all the poloxamer is completely dissolved.

Two mL of the above sample is placed in a 3 mL screw cap glass vial(with rubber lining) and closed tightly. The vial is placed in a MarketForge-sterilmatic autoclave (settings, slow liquids) and sterilized at250° F. for 25 minutes. After autoclaving the sample is left to cooldown to room temperature and then placed in refrigerator. The sample ishomogenized by mixing while the vials are cold.

Precipitation or discoloration are observed after autoclaving. HPLCanalysis is performed using an Agilent 1200 equipped with a Luna C18(2)3 μm, 100 Å, 250×4.6 mm column) using a 30-80 acetonitrile gradient(1-10 min) of (water-acetonitrile mixture containing 0.05% TFA), for atotal run of 15 minutes. Samples are diluted by taking 30 μL of sampleand dissolving with 1.5 mL of a 1:1 acetonitrile water mixture. Purityof the otic agent in the autoclaved samples is recorded.

Viscosity measurements are performed using a Brookfield viscometerRVDV-II+P with a CPE-51 spindle rotated at 0.08 rpm (shear rate of 0.31s⁻¹), equipped with a water jacketed temperature control unit(temperature ramped from 15-34° C. at 1.6° C./min). Tgel is defined asthe inflection point of the curve where the increase in viscosity occursdue to the sol-gel transition.

Dissolution is performed at 37° C. for the non-autoclaved sample insnapwells (6.5 mm diameter polycarbonate membrane with a pore size of0.4 μm), 0.2 mL of gel is placed into snapwell and left to harden, then0.5 mL is placed into reservoir and shaken using a Labline orbit shakerat 70 rpm. Samples are taken every hour (0.1 mL withdrawn and replacedwith warm buffer). Samples are analyzed for otic agent concentration byUV at 245 nm, against an external calibration standard curve.

Formulations comprising conivaptan, SR-121463 or micronized lixivaptanare tested using the above procedure to determine the effect addition ofa secondary polymer on the degradation products and viscosity of aformulation containing 2% otic agent and 17% poloxamer 407NF after heatsterilization (autoclaving).

Example 16 Effect of buffer type on the degradation products forformulations containing poloxamer 407NF after heat sterilization(autoclaving)

A TRIS buffer is made by dissolving 377.8 mg of sodium chloride (FisherScientific), and 602.9 mg of Tromethamine (Sigma Chemical Co.) then QSto 100 g with sterile filtered DI water, pH is adjusted to 7.4 with 1MHCl.

Stock solution containing 25% Poloxamer 407 solution in TRIS buffer:

Weigh 45 g of TRIS buffer, chill in an ice chilled bath then sprinkleinto the buffer, while mixing, 15 g of poloxamer 407 NF (SpectrumChemicals). The mixture is further mixed until all the poloxamer iscompletely dissolved.

A series of formulations is prepared with the above stock solution. Anappropriate amount of otic agent (or salt or prodrug thereof) and/orotic agent as micronized/coated/liposomal particles (or salt or prodrugthereof) is used for all experiments.

Stock solution (pH 7.3) containing 25% Poloxamer 407 solution in PBSbuffer:

PBS buffer described above is used. Dissolve 704 mg of sodium chloride(Fisher Scientific), 601.2 mg of sodium phosphate dibasic anhydrous(Fisher Scientific), 242.7 mg of sodium phosphate monobasic anhydrous(Fisher Scientific) with 140.4 g of sterile filtered DI water. Thesolution is cooled down in an ice chilled water bath and then 50 g ofpoloxamer 407NF (SPECTRUM CHEMICALS) is sprinkled into the cold solutionwhile mixing. The mixture is further mixed until the poloxamer iscompletely dissolved.

A series of formulations is prepared with the above stock solution. Anappropriate amount of otic agent (or salt or prodrug thereof) and/orotic agent as micronized/coated/liposomal particles (or salt or prodrugthereof) is used for all experiments.

Tables 2 and 3 list samples prepared using the procedures describedabove. An appropriate amount of otic agent is added to each sample toprovide a final concentration of 2% otic agent in the sample.

TABLE 2 Preparation of samples containing TRIS buffer 25% Stock TRISBuffer Sample pH Solution (g) (g) 20% P407/2 otic agent/TRIS 7.45 8.011.82 18% P407/2 otic agent/TRIS 7.45 7.22 2.61 16% P407/2 oticagent/TRIS 7.45 6.47 3.42 18% P4072 otic agent/TRIS 7.4 7.18 2.64  4%otic agent/TRIS 7.5 — 9.7  2% otic agent/TRIS 7.43 — 5  1% oticagent/TRIS 7.35 — 5  2% otic agent/TRIS (suspension) 7.4 — 4.9

TABLE 3 Preparation of samples containing PBS buffer (pH of 7.3) 25%Stock Solution Sample in PBS (g) PBS Buffer (g) 20% P407/2 oticagent/PBS 8.03 1.82 18% P407/2 otic agent/PBS 7.1 2.63 16% P407/2 oticagent/PBS 6.45 3.44 18% P407/2 otic agent/PBS — 2.63  2% otic agent/PBS— 4.9

One mL samples are individually placed in 3 mL screw cap glass vials(with rubber lining) and closed tightly. The vials are placed in aMarket Forge-sterilmatic autoclave (setting, slow liquids) andsterilized at 250° F. for 25 minutes. After the autoclaving the samplesare left to cool down to room temperature. The vials are placed in therefrigerator and mixed while cold to homogenize the samples.

HPLC analysis is performed using an Agilent 1200 equipped with a LunaC18(2) 3 μm, 100 Å, 250×4.6 mm column) using a 30-80 acetonitrilegradient (1-10 min) of (water-acetonitrile mixture containing 0.05%TFA), for a total run of 15 minutes. Samples are diluted by taking 30 μLof sample and dissolving with 1.5 mL of a 1:1 acetonitrile watermixture. Purity of the otic agent in the autoclaved samples is recorded.The stability of formulations in TRIS and PBS buffers is compared.

Viscosity measurements are performed using a Brookfield viscometerRVDV-II+P with a CPE-51 spindle rotated at 0.08 rpm (shear rate of 0.31s⁻¹), equipped with a water jacketed temperature control unit(temperature ramped from 15-34° C. at 1.6° C./min). Tgel is defined asthe inflection point of the curve where the increase in viscosity occursdue to the sol-gel transition. Only formulations that show no changeafter autoclaving are analyzed.

Formulations comprising conivaptan, SR-121463 or micronized lixivaptanare tested using the above procedure to determine the effect addition ofa secondary polymer on the degradation products and viscosity of aformulation containing 2% otic agent and 17% poloxamer 407NF after heatsterilization (autoclaving). Stability of formulations containingmicronized otic agent is compared to non-micronized otic agentformulation counterparts.

Example 17 Pulsed release otic formulations

A combination of SR-121463 and SR-121463 hydrochloride (ratio of 1:1) isused to prepare a pulsed release otic agent formulation using theprocedures described herein. 20% of the delivered dose of SR-121463 issolubilized in a 17% poloxamer solution of example 13 with the aid ofbeta-cyclodextrins. The remaining 80% of the otic agent is then added tothe mixture and the final formulation is prepared using any proceduredescribed herein.

Pulsed release formulations comprising conivaptan, SR-121463 ormicronized lixivaptan prepared according to the procedures and examplesdescribed herein, are tested using procedures described herein todetermine pulse release profiles.

Example 18 Preparation of a 17% poloxamer 407/2% otic agent/78 Ppm Evansblue in PBS

A Stock solution of Evans Blue (5.9 mg/mL) in PBS buffer is prepared bydissolving 5.9 mg of Evans Blue (Sigma Chemical Co) with 1 mL of PBSbuffer (from example 13).

A Stock solution containing 25% Poloxamer 407 solution in PBS buffer isused in this study. An appropriate amount of an otic agent is added tothe stock solution to prepare formulations comprising 2% of an oticagent (Table 4).

TABLE 4 Preparation of poloxamer 407 samples containing Evans Blue 25%P407in PBS Buffer Evans Blue Sample ID PBS (g) (g) Solution (μL) 17%P407/2 otic agent/EB 13.6 6 265 20% P407/2 otic agent/EB 16.019 3.62 26525% P407/2 otic agent/EB 19.63 — 265

Formulations comprising conivaptan, SR-121463 or micronized lixivaptanare prepared according to the procedures described above and are sterilefiltered through 0.22 μm PVDF syringe filters (Millipore corporation),and autoclaved.

The above formulations are dosed to guinea pigs in the middle ear byprocedures described herein and the ability of formulations to gel uponcontact and the location of the gel is identified after dosing and at 24hours after dosing.

Example 19 Terminal sterilization of poloxamer 407 formulations with andwithout a visualization dye

17% poloxamer 407/2% otic agent/in phosphate buffer, pH 7.3: Dissolve709 mg of sodium chloride (Fisher Scientific), 742 mg of sodiumphosphate dibasic dehydrate USP (Fisher Scientific), 251.1 mg of sodiumphosphate monobasic monohydrate USP (Fisher Scientific) and anappropriate amount of an otic agent with 158.1 g of sterile filtered DIwater. The solution is cooled down in an ice chilled water bath and then34.13 g of poloxamer 407NF (Spectrum chemicals) is sprinkled into thecold solution while mixing. The mixture is further mixed until thepoloxamer is completely dissolved.

17% poloxamer 407/2% otic agent/59 ppm Evans blue in phosphate buffer:Take two mL of the 17% poloxamer 407/2% otic agent/in phosphate buffersolution and add 2 mL of a 5.9 mg/mL Evans blue (Sigma-Aldrich chemicalCo) solution in PBS buffer.

25% poloxamer 407/2% otic agent/in phosphate buffer: Dissolve 330.5 mgof sodium chloride (Fisher Scientific), 334.5 mg of sodium phosphatedibasic dehydrate USP (Fisher Scientific), 125.9 mg of sodium phosphatemonobasic monohydrate USP (Fisher Scientific) and an appropriate amountof an otic agent with 70.5 g of sterile filtered DI water.

The solution is cooled down in an ice chilled water bath and then 25.1 gof poloxamer 407NF (Spectrum chemicals) is sprinkled into the coldsolution while mixing. The mixture is further mixed until the poloxameris completely dissolved.

25% poloxamer 407/2% otic agent/59 ppm Evans blue in phosphate buffer:Take two mL of the 25% poloxamer 407/2% otic agent/in phosphate buffersolution and add 2 mL of a 5.9 mg/mL Evans blue (Sigma-Aldrich chemicalCo) solution in PBS buffer.

Place 2 mL of formulation into a 2 mL glass vial (Wheaton serum glassvial) and seal with 13 mm butyl str (kimble stoppers) and crimp with a13 mm aluminum seal. The vials are placed in a Market Forge-sterilmaticautoclave (settings, slow liquids) and sterilized at 250° F. for 25minutes. After the autoclaving the samples are left to cool down to roomtemperature and then placed in refrigeration. The vials are placed inthe refrigerator and mixed while cold to homogenize the samples. Samplediscoloration or precipitation after autoclaving is recorded.

HPLC analysis is performed using an Agilent 1200 equipped with a LunaC18(2) 3 μm, 100 Å, 250×4.6 mm column) using a 30-95 methanol:acetatebuffer pH 4 gradient (1-6 min), then isocratic for 11 minutes, for atotal run of 22 minutes. Samples are diluted by taking 30 μL of sampleand dissolved with 0.97 mL of water. The main peaks are recorded in thetable below. Purity before autoclaving is always greater than 99% usingthis method.

Viscosity measurements are performed using a Brookfield viscometerRVDV-II+P with a CPE-51 spindle rotated at 0.08 rpm (shear rate of 0.31s⁻¹), equipped with a water jacketed temperature control unit(temperature ramped from 15-34° C. at 1.6° C./min). Tgel is defined asthe inflection point of the curve where the increase in viscosity occursdue to the sol-gel transition.

Formulations comprising conivaptan, SR-121463 or micronized lixivaptanprepared according to the procedures described herein, are tested usingthe above procedures to determine stability of the formulations.

Example 20 In vitro comparison of relase profile

Dissolution is performed at 37° C. in snapwells (6.5 mm diameterpolycarbonate membrane with a pore size of 0.4 μm), 0.2 mL of a gelformulation described herein is placed into snapwell and left to harden,then 0.5 mL buffer is placed into reservoir and shaken using a Lablineorbit shaker at 70 rpm. Samples are taken every hour (0.1 mL withdrawnand replace with warm buffer). Samples are analyzed for otic agentconcentration by UV at 245 nm against an external calibration standardcurve. Pluronic concentration is analyzed at 624 nm using the cobaltthiocyanate method. Relative rank-order of mean dissolution time (MDT)as a function of % P407 is determined. A linear relationship between theformulations mean dissolution time (MDT) and the P407 concentrationindicates that the otic agent is released due to the erosion of thepolymer gel (poloxamer) and not via diffusion. A non-linear relationshipindicates release of otic agent via a combination of diffusion and/orpolymer gel degradation.

Alternatively, samples are analyzed using the method described by LiXin-Yu paper [Acta Pharmaceutica Sinica 2008, 43 (2):208-203] andRank-order of mean dissolution time (MDT) as a function of % P407 isdetermined.

Formulations comprising conivaptan, SR-121463 or micronized lixivaptan,prepared according to the procedures described herein, are tested usingthe above procedure to determine the release profile of the otic agents.

Example 21 In vitro comparison of gelation temperature

The effect of Poloxamer 188 and an otic agent on the gelationtemperature and viscosity of Poloxamer 407 formulations is evaluatedwith the purpose of manipulating the gelation temperature.

A 25% Poloxamer 407 stock solution in PBS buffer and the PBS solutiondescribed above are used. Poloxamer 188NF from BASF is used. Anappropriate amount of otic agent is added to the solutions described inTable 5 to provide a 2% formulation of the otic agent.

TABLE 5 Preparation of samples containing poloxamer 407/poloxamer 18825% P407 Stock Poloxamer 188 PBS Buffer Sample Solution (g) (mg) (g) 16%P407/10% P188 3.207 501 1.3036 17% P407/10% P188 3.4089 500 1.1056 18%P407/10% P188 3.6156 502 0.9072 19% P407/10% P188 3.8183 500 0.7050 20%P407/10% P188 4.008 501 0.5032 20% P407/5% P188 4.01 256 0.770

Mean dissolution time, viscosity and gel temperature of the aboveformulations are measured using procedures described herein.

An equation is fitted to the data obtained and can be utilized toestimate the gelation temperature of F127/F68 mixtures (for 17-20% F127and 0-10% F68).T _(gel)=−1.8(% F127)+1.3(% F68)+53

An equation is fitted to the data obtained and can be utilized toestimate the Mean Dissolution Time (hr) based on the gelationtemperature of F127/F68 mixtures (for 17-25% F127 and 0-10% F68), usingresults obtained in examples above.MDT=−0.2(T _(gel))+8

Formulations comprising conivaptan, SR-121463 or micronized lixivaptanare prepared by addition of an appropriate amount of otic agents to thesolutions described in Table 5. The gel temperature of the formulationsis determined using the procedure described above.

Example 22 Determination of temperature range for sterile filtration

The viscosity at low temperatures is measured to help guide thetemperature range at which the sterile filtration needs to occur toreduce the possibility of clogging.

Viscosity measurements are performed using a Brookfield viscometerRVDV-II+P with a CPE-40 spindle rotated at 1, 5 and 10 rpm (shear rateof 7.5, 37.5 and 75 s⁻¹), equipped with a water jacketed temperaturecontrol unit (temperature ramped from 10-25° C. at 1.6° C./min).

The Tgel of a 17% Pluronic P407 is determined as a function ofincreasing concentration of otic agent. The increase in Tgel for a 17%pluronic formulation is estimated by:ΔT_(gel)=0.93[% otic agent]

Formulations comprising conivaptan, SR-121463 or micronized lixivaptan,prepared according to procedures described herein, are tested using theabove procedure to determine the temperature range for sterilefiltration. The effect of addition of increased amounts of otic agent onthe Tgel, and the apparent viscosity of the formulations is recorded.

Example 23 Determination of manufacturing conditions

TABLE 6 Viscosity of potential formulations at manufacturing/filtrationconditions. Apparent Viscosity^(a) (cP) Temperature Sample 5° C. belowTgel 20° C. @ 100 cP Placebo 52 cP @ 17° C. 120 cP   19° C. 17% P407/2%otic agent 90 cP @ 18° C. 147 cP 18.5° C. 17% P407/6% otic agent 142 cP@ 22° C.  105 cP 19.7° C. ^(a)Viscosity measured at a shear rate of 37.5s⁻¹

An 8 liter batch of a 17% P407 placebo is manufactured to evaluate themanufacturing/filtration conditions. The placebo is manufactured byplacing 6.4 liters of DI water in a 3 gallon SS pressure vessel, andleft to cool down in the refrigerator overnight. The following morningthe tank is taken out (water temperature 5° C., RT 18° C.) and 48 g ofsodium chloride, 29.6 g of sodium phosphate dibasic dehydrate and 10 gof sodium phosphate monobasic monohydrate is added and dissolved with anoverhead mixer (IKA RW20 @ 1720 rpm). Half hour later, once the bufferis dissolved (solution temperature 8° C., RT 18° C.), 1.36 kg ofpoloxamer 407 NF (spectrum chemicals) is slowly sprinkled into thebuffer solution in a 15 minute interval (solution temperature 12° C., RT18° C.), then speed is increased to 2430 rpm. After an additional onehour mixing, mixing speed is reduced to 1062 rpm (complete dissolution).

The temperature of the room is maintained below 25° C. to retain thetemperature of the solution at below 19° C. The temperature of thesolution is maintained at below 19° C. up to 3 hours of the initiationof the manufacturing, without the need to chill/cool the container.

Three different Sartoscale (Sartorius Stedim) filters with a surfacearea of 17.3 cm² are evaluated at 20 psi and 14° C. of solution

-   -   1) Sartopore 2, 0.2 μm 5445307HS-FF (PES), flow rate of 16        mL/min    -   2) Sartobran P, 0.2 μm 5235307HS-FF (cellulose ester), flow rate        of 12 mL/min    -   3) Sartopore 2 XLI, 0.2 μm 5445307IS-FF (PES), flow rate of 15        mL/min

Sartopore 2 filter 5441307H4-SS is used, filtration is carried out atthe solution temperature using a 0.45, 0.2 μm Sartopore 2 150 sterilecapsule (Sartorius Stedim) with a surface area of 0.015 m² at a pressureof 16 psi. Flow rate is measured at approximately 100 mL/min at 16 psi,with no change in flow rate while the temperature is maintained in the6.5-14° C. range. Decreasing pressure and increasing temperature of thesolution causes a decrease in flow rate due to an increase in theviscosity of the solution. Discoloration of the solution is monitoredduring the process.

TABLE 7 Predicted filtration time for a 17% poloxamer 407 placebo at asolution temperature range of 6.5-14° C. using Sartopore 2, 0.2 μmfilters at a pressure of 16 psi of pressure. Estimated flow rate Time tofilter 8 L Filter Size (m²) (mL/min) (estimated) Sartopore 2, size 40.015 100 mL/min 80 min Sartopore 2, size 7 0.05 330 mL/min 24 minSartopore 2, size 8 0.1 670 mL/min 12 min

Viscosity, Tgel and UV/Vis absorption is check before filtrationevaluation. Pluronic UV/Vis spectra are obtained by a Evolution 160UV/Vis (Thermo Scientific). A peak in the range of 250-300 nm isattributed to BHT stabilizer present in the raw material (poloxamer).Table 8 lists physicochemical properties of the above solutions beforeand after filtration.

TABLE 8 Physicochemical properties of 17% poloxamer 407 placebo solutionbefore and after filtration Viscosity^(a) @ Sample Tgel (° C.) 19° C.(cP) Absorbance @ 274 nm Before filtration 22 100 0.3181 Afterfiltration 22 100 0.3081 ^(a)Viscosity measured at a shear rate of 37.5s⁻¹

The above process is applicable for manufacture of 17% P407formulations, and includes temperature analysis of the room conditions.Preferably, a maximum temperature of 19° C. reduces cost of cooling thecontainer during manufacturing. In some instances, a jacketed containeris used to further control the temperature of the solution to easemanufacturing concerns.

Example 24 In vitro release of otic agent from an autoclaved micronizedsample

17% poloxamer 407/1.5% otic agent in TRIS buffer: 250.8 mg of sodiumchloride (Fisher Scientific), and 302.4 mg of Tromethamine (SigmaChemical Co.) is dissolved in 39.3 g of sterile filtered DI water, pH isadjusted to 7.4 with 1M HCl. 4.9 g of the above solution is used and anappropriate amount of micronized otic agent is suspended and dispersedwell. 2 mL of the formulation is transferred into a 2 mL glass vial(Wheaton serum glass vial) and sealed with 13 mm butyl styrene (kimblestoppers) and crimped with a 13 mm aluminum seal. The vial is placed ina Market Forge-sterilmatic autoclave (settings, slow liquids) andsterilized at 250° F. for 25 minutes. After the autoclaving the sampleis left to cool down to room temperature. The vial is placed in therefrigerator and mixed while cold to homogenize the sample. Samplediscoloration or precipitation after autoclaving is recorded.

Dissolution is performed at 37° C. in snapwells (6.5 mm diameterpolycarbonate membrane with a pore size of 0.4 μm), 0.2 mL of gel isplaced into snapwell and left to harden, then 0.5 mL PBS buffer isplaced into reservoir and shaken using a Labline orbit shaker at 70 rpm.Samples are taken every hour [0.1 mL withdrawn and replaced with warmPBS buffer containing 2% PEG-40 hydrogenated castor oil (BASF) toenhance otic agent solubility]. Samples are analyzed for otic agentconcentration by UV at 245 nm against an external calibration standardcurve. The release rate is compared to other formulations disclosedherein. MDT time is calculated for each sample.

Solubilization of otic agent in the 17% poloxamer system is evaluated bymeasuring the concentration of the otic agent in the supernatant aftercentrifuging samples at 15,000 rpm for 10 minutes using an eppendorfcentrifuge 5424. Otic agent concentration in the supernatant is measuredby UV at 245 nm against an external calibration standard curve.

Formulations comprising conivaptan, SR-121463 or micronized lixivaptan,prepared according to the procedures described herein, are tested usingthe above procedures to determine release rate of the otic agent fromeach formulation.

Example 25 Release rate or MDT and viscosity of formulation containingsodium carboxymethyl cellulose

17% poloxamer 407/2% otic agent/1% CMC (Hercules Blanose 7M): A sodiumcarboxymethylcellulose (CMC) solution (pH 7.0) in PBS buffer is preparedby dissolving 205.6 mg of sodium chloride (Fisher Scientific), 372.1 mgof sodium phosphate dibasic dihydrate (Fisher Scientific), 106.2 mg ofsodium phosphate monobasic monohydrate (Fisher Scientific) in 78.1 g ofsterile filtered DI water. 1 g of Blanose 7M CMC (Hercules, viscosity of533 cP @ 2%) is sprinkled into the buffer solution and heated to easesolution, solution is then cooled down and 17.08 g poloxamer 407NF(Spectrum Chemicals) is sprinkled into the cold solution while mixing. Aformulation comprising 17% poloxamer 407NF/1% CMC/2% otic agent in PBSbuffer is made adding/dissolving an appropriate amount of otic agent to9.8 g of the above solution, and mixing until all the otic agent iscompletely dissolved.

17% poloxamer 407/2% otic agent/0.5% CMC (Blanose 7M65): A sodiumcarboxymethylcellulose (CMC) solution (pH 7.2) in PBS buffer is preparedby dissolving 257 mg of sodium chloride (Fisher Scientific), 375 mg ofsodium phosphate dibasic dihydrate (Fisher Scientific), 108 mg of sodiumphosphate monobasic monohydrate (Fisher Scientific) in 78.7 g of sterilefiltered DI water. 0.502 g of Blanose 7M65 CMC (Hercules, viscosity of5450 cP @ 2%) is sprinkled into the buffer solution and heated to easesolution, solution is then cooled down and 17.06 g poloxamer 407NF(Spectrum Chemicals) is sprinkled into the cold solution while mixing. A17% poloxamer 407NF/1% CMC/2% otic agent solution in PBS buffer is madeadding/dissolving an appropriate amount of otic agent to 9.8 g of theabove solution, and mixing until the otic agent is completely dissolved.

17% poloxamer 407/2% otic agent/0.5% CMC (Blanose 7H9): A sodiumcarboxymethylcellulose (CMC) solution (pH 7.3) in PBS buffer is preparedby dissolving 256.5 mg of sodium chloride (Fisher Scientific), 374 mg ofsodium phosphate dibasic dihydrate (Fisher Scientific), 107 mg of sodiumphosphate monobasic monohydrate (Fisher Scientific) in 78.6 g of sterilefiltered DI water, then 0.502 g of Blanose 7H9CMC (Hercules, viscosityof 5600 cP @1%) is sprinkled to the buffer solution and heated to easesolution, solution is then cooled down and 17.03 g poloxamer 407NF(Spectrum Chemicals) is sprinkled into the cold solution while mixing. A17% poloxamer 407NF/1% CMC/2% otic agent solution in PBS buffer is madeadding/dissolving an appropriate amount of otic agent to 9.8 of theabove solution, and mixing until the otic agent is completely dissolved.

Viscosity measurements are performed using a Brookfield viscometerRVDV-II+P with a CPE-40 spindle rotated at 0.08 rpm (shear rate of 0.6s⁻¹), equipped with a water jacketed temperature control unit(temperature ramped from 10-34° C. at 1.6° C./min). Tgel is defined asthe inflection point of the curve where the increase in viscosity occursdue to the sol-gel transition.

Dissolution is performed at 37° C. in snapwells (6.5 mm diameterpolycarbonate membrane with a pore size of 0.4 μm). 0.2 mL of gel isplaced into snapwell and left to harden, then 0.5 mL PBS buffer isplaced into reservoir and shaken using a Labline orbit shaker at 70 rpm.Samples are taken every hour, 0.1 mL withdrawn and replaced with warmPBS buffer. Samples are analyzed for otic agent concentration by UV at245 nm against an external calibration standard curve. The release rateis compared to the formulations disclosed in above examples, and MDTtime is calculated for each of the above formulations.

Formulations comprising conivaptan, SR-121463 or micronized lixivaptan,prepared according to procedures described above, are tested using theabove procedures to determine relationship between release rate and/ormean dissolution time and viscosity of formulation containing sodiumcarboxymethyl cellulose. Any correlation between the mean dissolutiontime (MDT) and the apparent viscosity (measured at 2° C. below thegelation temperature) is recorded.

Example 26 Application of an Enhanced Viscosity Aural pressuremodulating agent formulation onto the round window membrane

A formulation according to Example 2 is prepared and loaded into 5 mlsiliconized glass syringes attached to a 15-gauge luer lock disposableneedle. Lidocaine is topically applied to the tympanic membrane, and asmall incision made to allow visualization into the middle ear cavity.The needle tip is guided into place over the round window membrane, andthe aural pressure modulator formulation applied directly onto theround-window membrane.

Example 27 In Vivo testing of Intratympanic Injection of aural pressuremodulating agent formulation in a guinea pig

A cohort of 21 guinea pigs (Charles River, females weighing 200-300 g)is intratympanically injected with 50 μL of different P407-DSPformulations described herein, containing 0 to 50% otic agent. The gelelimination time course for each formulation is determined. A faster gelelimination time course of a formulation indicates lower meandissolution time (MDT). Thus the injection volume and the concentrationof an aural pressure modulator in a formulation are tested to determineoptimal parameters for preclinical and clinical studies.

Example 28 In vivo extended release kinetics

A cohort of 21 guinea pigs (Charles River, females weighing 200-300 g)is intratympanically injected with 50 μL 17% Pluronic F-127 formulationbuffered at 280 mOsm/kg and containing 1.5% to 35% aural pressuremodulating agent by weight of the formulation. Animals are dosed onday 1. The release profile for the formulations is determined based onanalysis of the perilymph.

Example 29 Evaluation of VP2 Antagonist Formulations in an EndolymphaticHydrops Animal Model

The following procedure is used to determine the efficacy of thethermoreversible gel formulation of lixivaptan as prepared in Example 1.

Materials and Methods

Thirty-five Hartley guinea pigs with a positive Preyer's reflex andweighing about 300 g are used. Five animals, which serve as controls(normal ear group), are fed for 5 weeks with neither operation nortreatment, and the remaining 30 serve as experimental animals. Allexperimental animals received electro-cauterization of the endolymphaticsac (Lee et al., Acta Otolaryngol. (1992) 112:658-666; Takeda et al.,Equilib. Res. (1993) 9:139-143). Four weeks after surgery, these animalsare divided into three groups of non-infusion hydropic ears,vehicle-treated hydropic ears and lixivaptan-treated hydropic ears,consisting of 10 animals each. The group of non-infusion hydropic earsreceive no treatment except for electro-cauterization of theendolymphatic sac. In the groups of vehicle-treated hydropic ears andlixivaptan-treated hydropic ears, the thermoreversible gel formulationis applied to the round window membrane. One week after administrationof the composition, all animals are sacrificed for assessment of thechanges of the endolymphatic space. All animals are left undisturbed andfreely moving in individual cages in a quiet room throughout the period,except during experimental procedures.

To assess the changes to the endolymphatic space, all animals aretranscardially perfused with physiological saline solution under deepanesthesia by a peritoneal injection of pentobarbital, and fixation isperformed with 10% formalin. The left temporal bones are removed andpostfixed in 10% formalin solution for 10 days or more. Thereafter, theyare decalcified with 5% trichloroacetic acid for 12 days and dehydratedin a graded ethanol series. They are embedded in paraffin and celloidin.The prepared blocks are cut horizontally into 6 μm sections. Thesections are stained with hematoxylin and eosin and observed under alight microscope. Quantitative assessment of changes of theendolymphatic space is performed according to the method of Takeda(Takeda et al., Hearing Res. (2003) 182:9-18).

Example 30 Evaluation of Lixivaptan Administration in Meniere's Patients

Study Objective

The primary objective of this study will be to assess the safety andefficacy of Lixivaptan (100 mg) in ameliorating Meniere's Disease inhuman subjects.

Methods

Study Design

This will be a phase 3, multicentre, double-blind, randomised,placebo-controlled, parallel group study comparing lixivaptanadministration (100 mg) to placebo in the treatment of endolymphatichydrops. Approximately 100 subjects will be enrolled in this study, andrandomised (1:1) to 1 of 2 treatment groups based on a randomisationsequence prepared by the sponsor. Each group will receive either 100 mglixivaptan+meclizine or meclizine treatment alone.

Subjects who do not complete the study will not be replaced. Allpatients will receive daily meclizine treatment for 8 weeks. Patientsreceiving the study drug (Lixivaptan 100 mg or matching placebo) will beadministered a gel formulation directly onto the subjects' round windowmembrane for 8 weeks. Each patient will receive a vestibular and hearingevaluation before each treatment with meclizine and the study drug.

While preferred embodiments of the present invention have been shown anddescribed herein, such embodiments are provided by way of example only.Various alternatives to the embodiments described herein are optionallyemployed in practicing the inventions. It is intended that the followingclaims define the scope of the invention and that methods and structureswithin the scope of these claims and their equivalents be coveredthereby.

We claim:
 1. A pharmaceutical composition for otic administrationcomprising: (i) a therapeutically effective amount of a multiparticulateaural pressure modulating agent, or pharmaceutically acceptable saltthereof; (ii) a copolymer of polyoxyethylene and polyoxypropylene in anamount sufficient to provide a gelation temperature of between about 19°C. and about 42° C., a gelation viscosity between about 15,000 cP andabout 1,000,000 cP; and a non-gelation viscosity that allows injectionwith a 18-31 gauge needle through the tympanic membrane on or near theround window membrane; and (iii) having an osmolarity of 100-500 mOsm/L;wherein the pharmaceutical composition for otic administration providesa sustained release of the aural pressure modulating agent to the innerear for a period of at least 5 days after a single administration;wherein the aural pressure modulating agent is a vasopressin receptormodulator, prostaglandin receptor modulator, estrogen-related receptorbeta modulator, a calcium channel blocker, or combinations thereof;provided that the aural pressure modulating agent is anon-corticosteroid aural pressure modulating agent.
 2. Thepharmaceutical composition for otic administration of claim 1, whereinthe pharmaceutical composition for otic administration is anauris-acceptable thermoreversible gel.
 3. The pharmaceutical compositionfor otic administration of claim 1, wherein the aural pressuremodulating agent is in the form of a neutral molecule, free acid, freebase, a salt, or a combination thereof.
 4. The pharmaceuticalcomposition for otic administration of claim 1, wherein the copolymer ofpolyoxyethylene and polyoxypropylene is poloxamer
 407. 5. Thepharmaceutical composition for otic administration of claim 1, whereinthe aural pressure modulating agent is a prostaglandin.
 6. Thepharmaceutical composition for otic administration of claim 1, whereinthe pharmaceutical composition for otic administration provides asustained release of the aural pressure modulating agent to the innerear for a period of at least 7 days after a single administration.